Patent Description:
Plasma medicine has qualified as a new scientific field after intense research effort in low-temperature or cold atmospheric plasma applications. It is known that cold atmospheric plasmas ("CAP") produce various chemically reactive species including reactive oxygen species (ROS) and reactive nitrogen species (RNS). CAP is a cocktail containing ROS and RNS in combination with transient electric fields, UV and charged species.

CAP has already been proven to be effective in wound healing, skin diseases, hospital hygiene, sterilization, antifungal treatments, dental care, and cosmetics targeted cell/tissue removal. One of the most recent applications of CAP is in cancer therapy. As a near-room temperature ionized gas, cold atmospheric plasma (CAP) has demonstrated its promising capability in cancer treatment by causing the selective death of cancer cells in vitro. See, <NPL>); <NPL>); <NPL>). The CAP treatment on several subcutaneous xenograft tumors and melanoma in mice has also demonstrated its potential clinical application. See, <NPL>); <NPL>).

Additionally, various experiments have been performed in connection with the effect of CAP on viruses. In <NPL>, the authors reported successful inactivation of adenovirus, a non-enveloped double stranded DNA virus, in a solution using a surface micro-discharge technology operating in air. In <NPL>, the authors reported on their investigation of the inactivation efficacy of Newcastle disease virus by non-thermal plasma-activated solutions. In <NPL>), the authors reported on their study of the effectiveness of a packed bed dielectric barrier discharge (DBD) NTP reactor to inactivate bacteriophage MS2 in aerosols. See also, <CIT>, entitled "Production of Immune-response Stimulating Aerosols by Non-thermal Plasma Treatment of Airborne Pathogens.

Several different systems and methods for performing Cold Atmospheric Plasma (CAP) treatment have been disclosed. For example, <CIT> discloses a two-electrode system for CAP treatement of cancer cells.

Another exemplary Cold Atmospheric Plasma system is disclosed in <CIT>. The disclosed system has two units, namely a Conversion Unit (CU) and a Cold Plasma Probe (CPP). The Conversion Unit is connected to high frequency electrosurgical generator (ESU) output and converts the ESU signal to a signal appropriate for performing cold atmospheric plasma procedures. The Cold Plasma Probe is connected to the Conversion Unit output. At the end of the Cold Plasma Probe cold plasma is produced and is thermally harmless to living tissue, i.e., it cannot cause burns to the tissue. This cold plasma, however, is deadly for cancer cells while leaving normal cells unaffected. The disclosed Cold Plasma Conversion Unit is unique in that it utilizes a high voltage transformer to up-convert the voltage (<NUM>-<NUM> kV), down-convert the frequency (<<NUM>), and down-convert the power (<<NUM> W) of the high-voltage output from an electrosurgical unit (<CIT>).

Additional research has shown that these CAP systems can be used to stimulate media, which then can be used for cancer treatment. For example, <CIT>, discloses a method for preparing a CAP stimulated media for use in cancer treatment. Another method for preparing CAP stimulated media is disclosed in <CIT>.

Further, various systems and methods for controlling gas flow and an integrated gas-assisted electrosurgical generator having a graphical user interface is disclosed in <CIT>, entitled "Electrosurgical Gas Control Module" and <CIT>, entitled "Gas Enhanced Electrosurgical Generator.

A variety of medical ventilator systems have been disclosed. Medical ventilators typically have a source of pressurized oxygen, which is fluidly connected to a patient through a conduit. For example, <CIT> discloses a medical system having a ventilator coupled to a breaching circuit. Some ventilator systems add means for monitoring patient data. For example, <CIT> discloses systems and methods for managing ventilation of a patient being ventilated by a medical ventilator, and in particular, for integrating oximeter data with the medical ventilator. Another example is <CIT>, which discloses a ventilator-extracorporeal membrane gas-exchange (ECGE) system. Yet another example is <CIT>, which discloses a medical ventilator with a pneumonia and pneumonia bacterial disease analysis function by using gas recognition.

Still other systems include means for supplying a medical gas with a ventilator. <CIT> discloses a method and device for supplying at least one medical gas to a patient receiving artificial respiration with the aid of a ventilator. A gas mixture provided by a respiratory gas flow of a ventilator and a medical gas added to the flow are supplied to a connecting piece, such as a Y-piece or Y-connector from which a patient feed line leads to the mechanically ventilated patient and from which a further line branches off. Via this further line at least the gas exhaled by the patient and the proportion of the respiratory gas introduced to the first line by the ventilator and the medical gas fed into the first line which have not been inhaled by the patient are discharged via the second line. For example, <CIT> discloses a ventilator for supplying a mixed gas of oxygen and a medical gas other than oxygen to a patient. Other relevant prior arts are disclosed in <CIT>, <CIT>, <CIT> and <CIT>.

In a preferred embodiment, the present invention is a system for using cold atmospheric plasma to treat respiratory infections or cancers of the respiratory system, and, in particular, to treat patients having COVID-<NUM>.

In a preferred embodiment, the present invention is a system for performing plasma treatment of respiratory infections. "Plasma treatment of respiratory infections" as used herein refers to the use of a plasma to generate reactive species to be delivered to a patient's respiratory system. The system has a source of a carrier gas, a humidifier connected to the source of a carrier gas, a source of a feed gas, a humidifier connected to the source of a feed gas, a plasma generator configured to plasmatize the carrier gas into a plasma, a mixer and a fluid delivery member connected to an output of the mixer for delivering reactive species generated in the mixer to a patient. The mixer has an interior chamber formed from a dielectric, an active electrode inside the interior chamber and connected to an electrical output of the plasma generator, and an outer electrode connected to a ground, wherein the mixer has a first fluid input port connected to the source of a carrier gas and a second fluid input connected to the source of a feed gas. The structure of the mixer forms a dielectric barrier discharge system for generating plasma. The carrier gas may comprise at least one of helium, argon, nitrogen and oxygen. The delivery member, for example, may be endobronchial tube, a nasal cannula or a face mask. The source of a feed gas comprises one of a ventilator and a continuous positive airway pressure device and may comprise a mixture of air and oxygen.

The plasma generator preferably operates with a frequency in the range of <NUM> to <NUM> and an output peak voltage in the range of 3kV to 6kV. In a preferred embodiment, the plasma generator generates electrical energy having a frequency within <NUM> of one of <NUM>, <NUM> and <NUM>. In another preferred embodiment, the plasma generator generates electrical energy having a frequency of <NUM>. The plasma generator may be a combination a high frequency electrosurgical generator and a low frequency converter. The plasma generator may have a power module, a CPU for controlling the power module, a memory connected to the CPU and a power supply connected to the CPU. Still further, the plasma generator may have a touchscreen display, a controller connected to the touchscreen display and a graphical user interface configured to display data on the touchscreen display and receive input from a user through the touch-screen display. The plasma generator further may have a gas module. The source of a carrier gas may be connected to the gas module and the gas module controls a flow of the carrier gas to the mixer. The first humidifier may be connected between the gas module and the mixer or may be connected between the gas module and the source of a carrier gas.

In a preferred embodiment, the first humidifier is configured to humidify a carrier gas flowing from the source of a carrier gas to at least <NUM>% humidity and the second humidifier is configured to humidify a feed gas flowing from the source of a feed gas to at least <NUM>% humidity. For example, the first humidifier is configured to humidify a carrier gas flowing from the source of a carrier gas to <NUM>% humidity and the second humidifier is configured to humidify a feed gas flowing from the source of a feed gas to at least <NUM>% humidity.

In another embodiment, the present invention is a system for performing plasma treatment of respiratory system. The system has an electrical energy generator configured to generate electrical energy to plasmatize a carrier gas into a plasma and a dielectric barrier discharge ("DBD") mixer. The DBD mixer has an interior chamber formed from a dielectric, the interior chamber having a first input configured to fluidly connect to a source of a humidified carrier gas, a second input configured to connect to a source of a humidified feed gas, and an output configured to connect to a delivery member, an active electrode inside the interior chamber and connected to an electrical output of the electrical energy generator, and an outer electrode connected to a ground. A plasma is generated in the interior chamber when electrical energy is supplied from the electrical energy generator to the interior electrode while both humidified feed gas and humidified carrier gas flow into the interior chamber. The system further may have a first humidifier fluidly connected to the first input of the chamber in the dielectric barrier discharge assembly and a second humidifier fluidly connected to the second input of the chamber in the dielectric barrier discharge assembly. Still further, the system may have a source of un-humidified helium fluidly connected to an input of the first humidifier and a source of un-humidified air fluidly connected to an input of the second humidifier.

Still other aspects, features, and advantages of the present invention are readily apparent from the following detailed description, simply by illustrating preferable embodiments and implementations. The present invention is also capable of other and different embodiments and its several details can be modified in various obvious respects, all without departing from the scope of the present invention. Accordingly, the drawings and descriptions are to be regarded as illustrative in nature and not as restrictive. Additional objects and advantages of the invention will be set forth in part in the description which follows and in part will be obvious from the description or may be learned by practice of the invention.

For a more complete understanding of the present invention and the advantages thereof, reference is now made to the following description and the accompanying drawings, in which:.

Cold atmospheric-pressure plasma (CAP) generates numerous reactive oxygen species (ROS) and reactive nitrogen species (RNS), such as hydroxyl radical (·OH), singlet oxygen (<NUM>O<NUM>), nitrogen ion (N<NUM>+), atomic oxygen (O), and, as well as electrons, ions, and photons. Maximum concentration of these species can be reached with optimal amount of humidity in the gas. CAP-generated ROS and RNS can form hydrogen peroxide (H<NUM>O<NUM>), nitrite (NO<NUM>-), nitrate (NO<NUM>-), peroxynitrite (ONOO-) when interacting with biological fluid. Reactive species and radicals in the plasma phase (· OH, N<NUM>+,<NUM>O<NUM>, O) are short-lived species, whereas H<NUM>O<NUM>, NO<NUM>-, NO<NUM>-, and ONOO- in aqueous phase are long-lived species. The long-lived species will further interact with intracellular species and metabolic pathways, inducing cell apoptosis. The present invention provides a system with which a cold atmospheric plasma (non-thermal plasma) can be generated and the reactive species can be delivered to patients across much greater distances than in prior systems. While prior systems typically are used at a distance of <NUM>-<NUM> from the target tissue, the present invention delivers plasma to the patient from a distance of greater than <NUM>.

A cold atmospheric plasma system for treatment of respiratory infections in accordance with a first preferred embodiment of the present invention is described with reference to <FIG>. In this embodiment, the carrier as is helium and the feed gas is air. A helium gas source <NUM> is split into two lines <NUM>, <NUM>, with each of the two lines controlled by a mass flow controller (MFC) <NUM>. Line <NUM> is fluidly connected to a first Humidifier <NUM>. The Helium gas flow (<NUM> to <NUM>/min) in line <NUM> is passed through an H<NUM>O filled container (Humidifier <NUM>) and then fed into a mixing chamber <NUM>. The helium gas flow in the line <NUM> is fed directly into the mixing chamber <NUM>. In this manner, with the mass flow controllers <NUM> on the lines <NUM>, <NUM> a relative H<NUM>O saturation in the gas exiting the chamber <NUM> can be adjusted. Adjustment of the gas flow in the two lines <NUM>, <NUM> makes the overall flow rate and humidity fine tuning of the gas flow exiting the chamber <NUM> possible. The humidity may be in the range of <NUM>%-<NUM>% with a preferred humidity of at least <NUM>%. The total helium flow in this embodiment could be varied from <NUM>/min to <NUM>/min in all cases. The humidity of Helium gas in the chamber <NUM> is measured via calibrated High-Accuracy Humidity and Temperature Meter <NUM>. The humidified helium gas from the chamber <NUM> is fed into an electrosurgical generator <NUM>, referred to herein as a "Cold Atmospheric Plasma (CAP) Generator. " A variety of electrosurgical generators are known in the art and could be used with the present invention. The gas being fed into the Cold Atmospheric Plasma (CAP) Generator <NUM> is referred to herein as the "carrier gas.

At the same time, an un-humidified air supply <NUM> (feed gas) is split into two lines <NUM>, <NUM>. Each line <NUM>, <NUM> is controlled by a mass flow controllers (MFC) <NUM>. Line <NUM> is fluidly connected to a second Humidifier <NUM>. The air gas flow in line <NUM> is passed through an H<NUM>O filled container (Humidifier <NUM>) and then is fed into a mixing chamber <NUM>. The air gas flow in the line <NUM> is fed directly into the mixing chamber <NUM>. In this manner, with the mass flow controllers <NUM> on the lines <NUM>, <NUM> a relative H<NUM>O saturation in the air feed exiting the chamber <NUM> can be adjusted. Adjustment of the air flow in the two lines <NUM>, <NUM> makes the overall flow rate and humidity fine tuning possible. The humidity may be in the range of <NUM>%-<NUM>% with a preferred humidity of at least <NUM>%. The humidity of air in the chamber <NUM> is measured via calibrated High-Accuracy Humidity and Temperature Meter <NUM>. The humidified air from the chamber <NUM> and oxygen from an oxygen supply <NUM> are provided to a respiratory delivery system <NUM>, such as a ventilator, CPAP machine, BIPAP machine, or other known respiratory deliver system. The respiratory delivery system <NUM> will mix, adjust and measure the pressure, flowrate, ratio and frequency of the patient inbreath of exhaust air, oxygen and CO2. The output of the respiratory delivery system <NUM> is connected to the CAP joint mixer, for example, via tubing <NUM> and a connector <NUM>. In testing, humidifying the helium flow to <NUM>% humidity and the air flow to <NUM>% humidity proved to be effective.

The output of the CAP generator <NUM> and the respiratory delivery system <NUM> are connected to a dielectric barrier discharge (DBD) assembly <NUM>, referred to herein as a "CAP joint mixer. " A ground cable <NUM> connects an outer electrode of the CAP joint mixer <NUM> to a ground in the CAP generator <NUM>. While the grounding cable <NUM> is shown separate from the tubing <NUM> in <FIG>, other arrangements are possible in which the ground cable <NUM> is combined, for example, in a harness with the tubing <NUM>. Due to the presence of the H<NUM>O, the ionization of Helium and H<NUM>O to He+ + e- chemical reaction will happen simultaneously. The cold plasma-generated reactive species (H2O2, NO2-, NO3-, ONOO-, and <NUM>-) are produced.

The output of the CAP joint mixer <NUM> is connected to a delivery member <NUM>, which, for example, may be an endobronchial tube, oxygen CPAP (continuous positive airway pressure), BIPAP (Bilevel Positive Airway Pressure), ventilator face mask, or nasal O<NUM> cannula <NUM> to deliver reactive species <NUM>, e.g., H2O2, NO2-, NO3-, ONOO-, and <NUM>-, generated by the system into the patient's respiratory system.

A preferred embodiment of a dielectric barrier discharge (DBD assembly or CAP Joint Mixer <NUM> is described with reference to <FIG>. The DBD assembly <NUM> has a first entry port <NUM> for receiving a flow of a first gas (e.g., a carrier gas), a second entry port <NUM> for receiving a second gas (e.g., a feed gas), and an exit port <NUM> through which gases and reactive species generated in the DBD assembly exit the assembly. The assembly has a primary housing <NUM> having a portion <NUM> forming a chamber 212a within the primary housing <NUM>. At least the portion <NUM> forming the chamber 212a is a dielectric material. In a preferred embodiment, the entire primary housing <NUM> is formed of a dielectric material, but other embodiments are possible wherein only a portion of the primary housing <NUM> including the portion <NUM> that forms the chamber 212a is formed of a dielectric material. In still other embodiments, a dielectric material separate from the primary housing <NUM> may surround the chamber 212a. The portion <NUM> of the primary housing <NUM> forms the exit port <NUM> to which a delivery member, such as an endobracheal tube or other type of tube, may be connected. The invention is not limited to any particular type of delivery member or connection between the delivery member and the exit port <NUM>. The primary housing <NUM> has a first neck or connector portion <NUM> forming a first input port <NUM> for receiving a first gas, which in this embodiment is the carrier gas (e.g., helium), and a channel leading to the chamber 212a. The interior of the neck portion <NUM> is threaded for receiving an interior electrode <NUM>. The primary housing <NUM> further has a second neck or connector <NUM> forming a second input port <NUM> for receiving a second gas, which in this embodiment is a feed gas (e.g., an air/oxygen mixture) and a channel 216a leading to the chamber 212a. A second outer electrode <NUM> made of a conductive material, e.g., copper, surrounds the exterior of the dielectric forming the chamber 212a. As shown in <FIG> and <FIG>, an outer insulating layer <NUM> covers the outer electrode <NUM>. The outer insulating layer <NUM> is not shown in <FIG>. As shown in <FIG>, the outer electrode <NUM> is connected to a ground. The housing <NUM> has a lip or ridge <NUM> abutting the outer electrode <NUM>. Within an upper portion of the lip or ridge <NUM> is a hole channel 218a, which allows for the outer electrode <NUM> to be connected to a ground wire <NUM> (see <FIG>) via connecting wire 220a.

The inner electrode <NUM> is made of a conductive material and has within it a channel <NUM> through which the first gas (a carrier gas) flows. The electrode <NUM> has a neck <NUM> that extends into the chamber 212a. The channel <NUM> extends through the neck <NUM> such that the first gas (the carrier gas) can flow into the chamber 212a. The exterior of the electrode <NUM> two threaded portions 230a, 230b and a lip or ridge <NUM>. The threaded portion 230a engages with the threaded interior of the neck <NUM> of the primary housing <NUM> to secure the inner electrode <NUM> into the primary housing <NUM>. The ridge or lip <NUM> of the electrode <NUM> provides a stop when the electrode <NUM> is fully threaded into the neck <NUM>.

The dielectric barrier discharge (DBD) assembly <NUM> further has a secondary housing <NUM> having a channel within it through which the first gas (the carrier gas) can flow to the channel <NUM> in the inner electrode <NUM>. The secondary housing <NUM> has a portion <NUM> having interior threaded for engaging the threaded portion 230b of the electrode <NUM> and thereby securing the secondary housing to the electrode <NUM> and the primary housing <NUM>. The secondary housing <NUM> has at the end of the threaded portion <NUM> a recess for receiving the electrode ridge or lip <NUM> and abutting the neck <NUM> of the primary housing <NUM>. The secondary housing <NUM> further has a connector structure <NUM>, 254a, 254b for connecting to a hose or other tubing <NUM> and connector <NUM> to connect the dielectric barrier discharge (DBD) assembly to the CAP generator <NUM>. Within the secondary housing <NUM> is a tube <NUM> through which the first gas (the carrier gas) flows. Within the tube <NUM> is an elongated electrode or wire that is connected to a conductive connector <NUM> (e.g., via solder). The conductive connector <NUM> abuts the inner electrode <NUM> and thus is electrically connected to the inner electrode <NUM>.

As shown in <FIG>, an exemplary cold atmospheric plasma (CAP) generator <NUM> has a power supply <NUM>, a CPU (or processor or FPGA) <NUM> and a memory or storage <NUM>. The system further has a display <NUM> (<FIG>), which may be the display of a tablet computer. The CPU <NUM> controls the system and receives input from a user through a graphical user interface displayed on display <NUM>. The CAP generator further has a gas control module <NUM> connected to a source <NUM> of a CAP carrier gas such as helium to control the flow of the carrier gas to the CAP joint mixer. The CAP generator <NUM> further has a radio frequency (RF) power module <NUM> for generating radio frequency (RF) energy. The RF power module contains conventional electronics such as are known for providing RF power in electrosurgical generators. The RF Power module operates with a frequency between <NUM>-<NUM> and output peak voltage from 3kV to 6kV and preferably at a frequency near (within <NUM>%) of <NUM>, <NUM> or <NUM>. The gas module <NUM> and RF power module <NUM> are connected to connector <NUM> that allows for CAP joint mixer <NUM> (or a CAP applicator <NUM> in FIGs. 11A and 11B) to be connected to the generator <NUM> via a connector having an electrical connector 196a and gas connector 196b.

As shown in <FIG>, other arrangements for delivery of the carrier gas and the electrical energy may be used with the invention. In <FIG>, a source <NUM> of a carrier gas (helium in this example) is provided to a gas control system <NUM> of any type, which supply the gas at a controlled flow rate to CAP joint mixer <NUM>. A conventional electrosurgical generator 350a supplies high frequency (HF) energy to a low frequency converter 350b, which outputs electrical energy having a frequency in the range of <NUM> to <NUM> and an output voltage in the range of 3kV to 6Kv.

Another embodiment, shown in <FIG>, has a carrier gas source <NUM> connected to a conventional gas control system <NUM>, which in turn is connected to the CAP joint mixer <NUM>, and a conventional electrosurgical generator <NUM> also connected to the CAP joint mixer <NUM>.

A generator housing <NUM> for a CAP-enabled gas-enhanced electrosurgical generator <NUM> in accordance with a preferred embodiment of the present invention is shown in <FIG>. The generator housing <NUM> has a housing <NUM> made of a sturdy material such as plastic or metal similar to materials used for housings of conventional electrosurgical generators. The housing <NUM> has a removable cover <NUM>. The housing <NUM> and cover <NUM> have means, such as screws, tongue and groove, or other structure for removably securing the cover to the housing. The cover <NUM> may comprise just the top of the housing or multiple sides, such as the top, right side, and left side, of the housing <NUM>. The housing <NUM> may have a plurality of feet or legs attached to the bottom of the housing. The bottom of the housing <NUM> may have a plurality of vents for venting from the interior of the gas-enhanced generator.

On the face of the housing <NUM> there is a touch-screen display <NUM> and a plurality of connectors <NUM>, <NUM> for connecting various accessories to the generator, such as an argon plasma probe, a hybrid plasma probe, a cold atmospheric plasma probe, or any other electrosurgical attachment. The face of the housing <NUM> is at an angle other than <NUM> degrees with respect to the top and bottom of the housing <NUM> to provide for easier viewing and use of the touch screen display <NUM> by a user. One or more of the gas control modules may be mounted within a gas-enhanced electrosurgical generator <NUM>.

The CAP-enabled gas-assisted electrosurgical generator has a graphical user interface (GUI) for controlling the components of the system using the touch screen display <NUM>. The graphical user interface for example, may control robotics, argon-monopolar cut/coag, hybrid plasma cut, cold atmospheric plasma, bipolar, plasma sealer, hemo dynamics or voice activation. The graphical user interface further may be used with fluorescence-guided surgery. The graphical user interface (GUI) further may be used with guided imaging such as CT, MRI, or ultrasound. The graphical user interface may communicate with RFID (such as may be found in various electrosurgical attachments) and may collect and store usage data in a storage medium. The graphical user interface communicates with the field-programmable gate array ("FPGA"), which may control an irrigation pump, insufflator, full bridge for adjusting the power output, fly back for regulating the power (DC to AC) and a foot pedal. The GUI further communicates with a database of data with associated predicted CAP settings or dosages via the CPU <NUM>. The database storage may be internal memory or other internal storage <NUM> or external storage.

A method for treatment of a respiratory infection in which both the feed gas (air) and the carrier gas (helium) are humidified is described with reference to <FIG>. Pressurized feed gas (air) is supplied to a humidifier <NUM>. The pressurized air is humidified in a humidifier <NUM>. Oxygen is added to the humidified air flow <NUM>. The humidified air and oxygen flow is controlled by a ventilator or other respiratory delivery system <NUM>. At the same time, a CAP carrier gas such as helium is supplied to a humidifier <NUM>. The carrier gas is humidified in the humidifier <NUM>. The humidified carrier gas is supplied to a CAP generator. The humidified CAP carrier gas from the CAP generator and the output of the ventilator both are supplied to a CAP joint mixer <NUM>, <NUM>. Electrical energy is applied to an inner electrode in the CAP joint mixer <NUM>. The output of the CAP joint mixer is then supplied to the patient's respiratory system <NUM>, for example, via a respiratory face mask, nasal cannula, or endobronchial tube.

In studies on the treatment of cancer using cold atmospheric plasma, it has been found that the CAP treatment decreases viability of cancer cells in a dose-dependent manner.

A method for treatment a respiratory infection in which both the feed gas (air) and the carrier gas (helium) are humidified is described with reference to <FIG>. In this embodiment, rather than electrical energy being applied to the inner electrode in the CAP joint mixer at a single setting (e.g., <NUM> V) for the entire treatment time (step <NUM> in <FIG>), the generator automatically sweeps thought a plurality of settings applying a first setting (e.g., <NUM> V) for a first time t<NUM> <NUM> while supplying the output to the patient <NUM> and then applying a second setting (e.g., <NUM> V) for a second time t<NUM> <NUM> while supplying the output to the patient <NUM>.

An alternative embodiment of a system <NUM> is referred to herein as the "Helium Gas Humidity Adjustment Setup" is described with reference to <FIG>. A helium gas source <NUM> is split into two lines <NUM>, <NUM>, with each of the two lines controlled by a mass flow controller (MFC) <NUM>. The Helium gas flow (<NUM> to <NUM>/min) in line <NUM> is passed through an H<NUM>O filled container (Humidifier <NUM>) and then is fed into a mixing chamber <NUM>. The helium gas flow in the line <NUM> is fed directly into the mixing chamber <NUM>. In this manner, with the mass flow controllers <NUM> on the lines <NUM>, <NUM> a relative H<NUM>O saturation in the gas exiting the chamber <NUM> can be adjusted. Adjustment of the gas flow in the two lines <NUM>, <NUM> makes the overall flow rate and humidity fine tuning of the gas flow exiting the chamber <NUM> possible. The humidity may be in the range of <NUM>%-<NUM>% with a preferred humidity of at least <NUM>%. The total helium flow in this embodiment could be varied from <NUM>/min to <NUM>/min in all cases. In this Helium Gas Humidity Adjustment Setup, the Helium H<NUM>O vapor content was varied during experiments. The humidity of Helium gas in the chamber <NUM> was measured via calibrated High-Accuracy Humidity and Temperature Meter <NUM> as shown in the <FIG>. The humidified helium gas from the chamber <NUM> is fed into a CAP generator <NUM>,.

At the same time, an un-humidified air tank <NUM> and un-humidified oxygen tank <NUM> feed air and oxygen respectively to a respiratory delivery system <NUM>, such as a ventilator, CPAP (continuous positive airway pressure) system, or BIPAP (Bilevel Positive Airway Pressure) system. These are referred to herein as "feed gases. " The respiratory delivery system <NUM> will mix, adjust, and measure the pressure, flow rate, ratio, and frequency of the patient inbreath of air, oxygen, and CO<NUM>. The output of the CAP generator <NUM> and the respiratory delivery system <NUM> are connected to a dielectric barrier discharge (DBD) assembly <NUM>. The output of the CAP joint mixer <NUM> is connected to a delivery member <NUM>, which, for example, may be an endobronchial tube, oxygen CPAP (continuous positive airway pressure), BIPAP (Bilevel Positive Airway Pressure), ventilator face mask, or nasal O<NUM> cannula <NUM> to deliver reactive species <NUM>, e.g., H2O2, NO2-, NO3-, ONOO-, and <NUM>-, generated by the system into the patient's respiratory system.

A method for treating a respiratory infection with the system of <FIG> is described with reference to <FIG>. A CAP carrier gas such as helium is supplied to a humidifier <NUM>. The carrier gas is humidified <NUM>. The humidified carrier gas is supplied to a CAP generator. At the same time, pressurized feed gas (air) is supplied <NUM>. Oxygen is added to the air flow <NUM>. The air and oxygen flow is controlled by a ventilator or other respiratory delivery system <NUM>. The humidified CAP carrier gas from the CAP generator and the output of the ventilator both are supplied to a CAP joint mixer <NUM>, <NUM>. Electrical energy is applied to an inner electrode in the CAP joint mixer <NUM>. The output of the CAP joint mixer is then supplied to the patient's respiratory system <NUM>, for example, via a respiratory face mask, nasal cannula, or endobronchial tube.

Another embodiment of a cold atmospheric plasma system <NUM> for treatment of respiratory infections is described with reference to <FIG>. A helium gas source <NUM> is fed into an electrosurgical generator <NUM>. At the same time, an un-humidified air supply <NUM> (feed gas) is split into two lines <NUM>, <NUM>. Each line <NUM>, <NUM> is controlled by a mass flow controllers (MFC) <NUM>. The air gas flow in line <NUM> is passed through an H<NUM>O filled container (Humidifier <NUM>) and then is fed into a mixing chamber <NUM>. The air gas flow in the line <NUM> is fed directly into the mixing chamber <NUM>. In this manner, with the mass flow controllers <NUM> on the lines <NUM>, <NUM> a relative H<NUM>O saturation in the air feed exiting the chamber <NUM> can be adjusted. Adjustment of the air flow in the two lines <NUM>, <NUM> makes the overall flow rate and humidity fine tuning possible. The humidity may be in the range of <NUM>%-<NUM>% with a preferred humidity of at least <NUM>%. The humidity of air in the chamber <NUM> is measured via calibrated High-Accuracy Humidity and Temperature Meter <NUM>. The humidified air from the chamber <NUM> and oxygen from an oxygen supply <NUM> are provided to a respiratory delivery system <NUM>, such as a ventilator, CPAP machine, BIPAP machine, or other known respiratory deliver system. The respiratory delivery system <NUM> will mix, adjust and measure the pressure, flowrate, ratio and frequency of the patient inbreath of exhaust air, oxygen and CO2.

A method for treatment a respiratory infection in which the feed gas (air) is humidified is described with reference to <FIG>. Pressurized feed gas (air) is supplied to a humidifier <NUM>. The pressurized air is humidified in a humidifier <NUM>. Oxygen is added to the humidified air flow <NUM>. The humidified air and oxygen flow is controlled by a ventilator or other respiratory delivery system <NUM>. At the same time, a CAP carrier gas such as helium is supplied to a CAP generator <NUM>. The CAP carrier gas from the CAP generator and the output of the ventilator both are supplied to a CAP joint mixer <NUM>, <NUM>. Electrical energy is applied to an inner electrode in the CAP joint mixer <NUM>. The output of the CAP joint mixer is then supplied to the patient's respiratory system <NUM>, for example, via a respiratory face mask, nasal cannula, or endobronchial tube. Other embodiments of the invention are possible in which the plasma is delivered to a patient, for example, through an endoscopic or laparoscopic device. Still further, in other embodiments, the present invention may treat cancer in the abdomen by feeding the output of the CAP joint mixer into the abdomen, for example, via a laparoscope or trocar.

A cold atmospheric plasma system for treatment of a patient via an endoscope or laparoscope in accordance with a preferred embodiment of the present invention is described with reference to <FIG>. A helium gas source <NUM> is split into two lines <NUM>, <NUM>, with each of the two lines controlled by a mass flow controller (MFC) <NUM>. The Helium gas flow (<NUM> to <NUM>/min) in line <NUM> is passed through an H<NUM>O filled container (Humidifier <NUM>) and then fed into a mixing chamber <NUM>. The helium gas flow in the line <NUM> is fed directly into the mixing chamber <NUM>. In this manner, with the mass flow controllers <NUM> on the lines <NUM>, <NUM> a relative H<NUM>O saturation in the gas exiting the chamber <NUM> can be adjusted. Adjustment of the gas flow in the two lines <NUM>, <NUM> makes the overall flow rate and humidity fine tuning of the gas flow exiting the chamber <NUM> possible. The humidity may be in the range of <NUM>%-<NUM>% with a preferred humidity of at least <NUM>%. The total helium flow in this embodiment could be varied from <NUM>/min to <NUM>/min in all cases. The humidity of Helium gas in the chamber <NUM> is measured via calibrated High-Accuracy Humidity and Temperature Meter <NUM>. The humidified helium gas from the chamber <NUM> is fed into an electrosurgical generator <NUM>. A variety of electrosurgical generators are known in the art and could be used with the present invention. The gas being fed into the Cold Atmospheric Plasma (CAP) Generator <NUM> is referred to herein as the "carrier gas.

At the same time, an un-humidified air supply <NUM> (feed gas) is split into two lines <NUM>, <NUM>. Each line <NUM>, <NUM> is controlled by a mass flow controllers (MFC) <NUM>. The air gas flow in line <NUM> is passed through an H<NUM>O filled container (Humidifier <NUM>) and then is fed into a mixing chamber <NUM>. The air gas flow in the line <NUM> is fed directly into the mixing chamber <NUM>. In this manner, with the mass flow controllers <NUM> on the lines <NUM>, <NUM> a relative H<NUM>O saturation in the air feed exiting the chamber <NUM> can be adjusted. Adjustment of the air flow in the two lines <NUM>, <NUM> makes the overall flow rate and humidity fine tuning possible. The humidity may be in the range of <NUM>%-<NUM>% with a preferred humidity of at least <NUM>%. The humidity of air in the chamber <NUM> is measured via calibrated High-Accuracy Humidity and Temperature Meter <NUM>. Humidified air from the chamber <NUM> and oxygen from an oxygen supply <NUM> are connected to a gas control system <NUM>. In an alternative embodiment, an integrated gas-enhanced electrosurgical generator having a plurality of gas control modules 1000a, 1000b, 1000c such as is shown in <FIG> may be used. In such a system the flow helium, air and oxygen all are controlled by gas modules in a single housing and having a unified control system.

The output of the CAP generator <NUM> and humidified air and oxygen from the gas control system <NUM> are connected to a dielectric barrier discharge (DBD) assembly <NUM>, referred to herein as a "CAP joint mixer. " A ground cable <NUM> connects an outer electrode of the CAP joint mixer <NUM> to a ground in the CAP generator <NUM>. While the grounding cable <NUM> is shown separate from the tubing <NUM> in <FIG>, other arrangements are possible in which the ground cable <NUM> is combined, for example, in a harness with the tubing <NUM>. Due to the presence of the H<NUM>O, the ionization of Helium and H<NUM>O to He+ + e- chemical reaction will happen simultaneously. The cold plasma-generated reactive species (H2O2, NO2-, NO3-, ONOO-, and <NUM>-) are produced.

The output of the CAP joint mixer <NUM> is connected to an elongated delivery member 190a, which, for example, may be a rigid or flexible tube of a size that will fit into a channel of any type of endoscope or laparoscope, whether the scope is a bronchoscope, colonoscope or any other type of scope used in surgical applications.

The embodiment shown in <FIG> where both the feed gas (air) and the carrier gas (helium) are humidified provides greater humidity in CAP joint mixer than embodiments where only one of the feed gas and carrier gas is humidified. In the embodiment of <FIG>, experiments have shown that due to the increased humidity, the treatment time necessary to achieve a <NUM>% kill rate for lung cancer cells can be reduced to <NUM> minutes versus about <NUM> minutes for the embodiments where only one of the feed gas and carrier gas is humidified. Further, the production of ozone can be reduced from roughly <NUM> ppm (parts per million) for the embodiments of <FIG> and <FIG> to less than <NUM> ppm (approximately <NUM> ppm) in the embodiment of <FIG>.

A gas control module <NUM> in accordance with the present invention is designed for gas-enhanced electrosurgical systems. Conventionally, gas-enhanced electrosurgical systems have an electrosurgical generator and a gas control unit that have separate housings. The conventional gas control unit typically controls only a single gas such as argon, CO<NUM> or helium. The present invention uses a gas control module <NUM> that may be used in a gas control unit or in a combined unit functioning both as an electrosurgical generator and as a gas control unit. Further, a plurality of gas control modules in accordance with the present invention may be combined in a single gas control unit or combination generator/gas control unit to provide control of multiple gases and provide control for multiple types of gas-enhanced surgery such as argon gas coagulation, hybrid plasma electrosurgical systems and cold atmospheric plasma systems.

Still further, while helium is the carrier gas used in the disclosed embodiments, other gases such as argon, nitrogen, oxygen or air may be used as a carrier gas.

While the preferred embodiments are described with a ventilator, other medical respiration devices such as a continuous positive airway pressure (CPAP) system could be used with the present invention.

The cold atmospheric plasma system for treatment of respiratory infections where only the air was humidified was used to treat <NUM> phosphate buffer saline (PBS) in <NUM>-well plates with the generator in Argon Coag Mode and Spray Mode operating at a frequency near <NUM> for <NUM> each. Voltage was set to be <NUM> V. Oxygen and air flow rate were both set to be <NUM> LPM. The mixture of oxygen (O<NUM>) and air was humidified by bubbling through DI water. The relative humidity (RH) of the mixture is about <NUM>%. Flow rate of helium, the carrier gas for cold atmospheric plasma (CAP), was set at <NUM> LPM. Therefore, the O<NUM> percentage of the final output gas from the endobronchial tube was about <NUM>% in the O<NUM>-air-He mixture.

Among of the cocktail of plasma-generated reactive oxygen species (ROS) and reactive nitrogen species (RNS) in the treated solution, hydrogen peroxide (H<NUM>O<NUM>) and nitrite (NO<NUM>-) are the most commonly studied long-lived species. Their concentrations were measured in treated phosphate-buffered saline (PBS) using Griess Reagent System (Promega, G2930) and colorimetric Hydrogen Peroxide Assay Kit (Sigma-Aldrich, MAK311-1KT) with CAP on or off. Results were read by a BioTek microplate reader at <NUM> and <NUM> for absorbance, respectively.

Electrosurgical generators typically have multiple modes of operation, including "cut" or cutting modes and "coag" or coagulation modes of operation. A cut mode typically will have a low voltage waveform form (e.g., 1KV) with a high duty cycle, e.g. <NUM>%. The coag mode of an electrosurgical generator typically creates a waveform with large amplitude but short duration "spikes" to achieve hemostasis (coagulation). For example, a coag mode on an electrosurgical generator may use a high voltage wave form at a <NUM>% duty cycle. Different degrees of hemostasis (coagulation) can be achieved by utilizing varying degrees of "Blended" waveforms, e.g., <NUM>% on/<NUM>% off, <NUM>% on/<NUM>% off, or <NUM>% on/<NUM>% off. Electrosurgical generators also have argon plasma coagulation modes, or "argon coag" modes. Argon Plasma Coagulation (APC) utilizes plasma produced by the ionization of a few millimeter diameter argon flow exhausting into ambient air from the electrosurgical hand-piece. When compared to a cut mode, an argon coagulation mode on a generator may use a high voltage (e.g. ,<NUM> KV for argon coag versus 1KV for a cut mode), less current (e.g., 200mA for argon coag versus 500mA for cut), and lower frequency (<NUM> for argon coag versus <NUM> for cut). Electrosurgical generators also have a "Spray Mode," which is similar to the argon coag mode (similar voltage and current), but they have a random week of frequency, for example, from <NUM>-<NUM>, which allows it to cover different tissue impedances.

The concentrations of H<NUM>O<NUM> and NO<NUM>- with the present system treatment were plotted in <FIG>. With CAP turned on, both species are higher when treated in Argon Coag Mode than when treated in Spray Mode. Gas-only treatment was also performed as a control. As indicated in <FIG>, with <NUM> of treatment, Argon Coag Mode at <NUM> V generated <NUM> H<NUM>O<NUM> and <NUM> NO<NUM>-; whereas Spray Mode at <NUM> V generated <NUM> H<NUM>O<NUM> and undetectable amount of NO<NUM>-. A gas mixture alone does not generate significant amount of ROS or RNS.

Cold atmospheric plasma has been reported to induce inactivation of airborne viruses (<NPL>)), deactivation of hepatitis B virus while keeping normal liver function during CAP treatment (<NPL>), inhibition of HIV replication (<NPL>), inactivation of Newcastle disease virus and avian influenza virus without destruction of the antigenic determinants for vaccine preparation (<NPL>) and so forth. In this study, CAP is combined with a ventilator system to achieve the delivery of CAP as well as the treatment of the virus throughout the patient's respiratory system.

Wu el al (<NPL>) suggested that the ambient air as carrier gas produced the highest level of inactivation at power levels of <NUM> and <NUM> W, followed by the gas carriers Ar-O<NUM> (<NUM>%, vol/vol) and He-O2 (<NUM>%, vol/vol). In addition, air is a required input gas for all ventilators. Hence air as carrier gas is the best option for a CAP-equipped ventilator. Relative humidity (RH) as an important factor of the air will be studied for an optimal configuration in addition to CAP treatment parameters including discharge voltage (V) and treatment time (t).

Reactive species generated by CAP can be confirmed within the plasma beam using optical emission spectroscopy (OES) and in aqueous solution by kits based on species.

An optical emission spectrometer (Ocean Optics HR2000) is used to detect the species in the plasma beam in the range of <NUM>-<NUM>. The plasma emissions are collected in a direction perpendicular to the plasma beam axis and at <NUM>-mm increments in axial direction using a collimating lens. The plasma emission is transmitted to the spectrometer via optical fiber.

Kondeti et al did a thorough research on the species generated in CAP-treated saline and water based on their half-lives. They concluded that long-lived species played a dominant role when the plasma was not in direct contact with the saline; whereas short-lived species was more important when the plasma touched the liquid. Long-lived species in CAP-treated solution like NO<NUM>- and H<NUM>O<NUM> concentrations can be measured in air flow-treated phosphate buffer saline (PBS) using Griess Reagent System (Promega, G2930) and Fluorimetric Hydrogen Peroxide Assay Kit (Sigma-Aldrich, MAK165-1KT) with CAP on or off. Results will be read by a BioTek microplate reader at <NUM> for absorbance and <NUM>/<NUM> for fluorescence, respectively.

Ozone can be a concern for CAP-based ventilator because it can have detrimental impacts on human health. Ozone concentration should be measured at the exhaust of the ventilator and reduced with filters to meet the air quality standards.

Lung cancer cell line A549 will be used for the efficacy of CAP treatment. Air flow-only treatment will be used as control group. The viability of the cells will be evaluated by <NUM>-(<NUM>,<NUM>-dimethylthiazol-<NUM>-yl)-<NUM>,<NUM>-diphenyl tetrazolium bromide (MTT) assay.

In conclusion, with tuned configuration, CAP-based ventilators could benefit patients with respiratory diseases like pneumonia, COVID19, or lung cancer.

Respiratory disease-causing viruses such as COVID-<NUM> and severe acute respiratory syndrome (SARS) are transmitted by aerosolized droplets containing the infection virus. Cold atmospheric-pressure plasma (CAP) generates numerous reactive oxygen species (ROS) and reactive nitrogen species (RNS), such as hydrogen peroxide (H<NUM>O<NUM>), singlet oxygen (<NUM>O2), ozone (O3), nitric oxide (·NO), and hydroxyl radical (·OH), as well as electrons, ions, and photons, in which ·OH, <NUM>O<NUM>, ·NO, O<NUM>·-, ·NO<NUM>, and ONOO- are short-lived species whereas H<NUM>O<NUM>, NO<NUM>-, and NO<NUM>-, are long-lived species. Various studies have shown that the CAP could inactivate viruses and other microbes. The potential inactivation mechanisms of the virus by the CAP is by inducing to high oxidation-reduction potential (ORP) and electrical conductivity by producing large number of free radicals. The reactive oxygen and nitrogen species could react with carbohydrates and initiate lipid peroxidation and cross-linking of the fatty acid side chains, resulting in alterations of the chemical bonds and molecular structure. They induce oxidative stress by causing protein peroxidation and inducing the destruction of the virus envelope, singlet oxygen could rapidly react with cysteine to generate the major product of cystine (R-cys-S-S-cys-R) with disulfides, they selectively reacted with tyrosine, tryptophan, and histidine to produce hydroperoxides resulting in protein aggregation and ultimately resulting in changes to the viral morphology. Moreover, they can damage viral nucleic acids encoding enzymes, by oxidizing guanine and induce cross-links between guanine and lysine contributing to reduced gene expression and the elimination of virus replication, thereby leading to virus inactivation. The cold plasma system of the present invention generates ionized cold plasma in a humidified setup to produce reactive species that are fed to virus infected patient via endobronchial tube. The output of the system contains reactive oxygen species (ROS) and reactive nitrogen species (RNS) which would inactivate the virus present in the patient's bronchial cells.

Human epithelial lung carcinoma cell line A549 (ATCC, CCL-<NUM>) was used to study the efficacy of the present invention. A549 cells were seeded at a density of <NUM><NUM>/well in <NUM>-well plates and treated for up to <NUM> minutes. The frequency was near <NUM>. Voltage was set to be <NUM> V. Flow rate of helium, the carrier gas for cold atmospheric plasma (CAP), was set at <NUM> LPM. The total flow rate of oxygen (O<NUM>) and air were set to be <NUM> LPM. The O<NUM> percentage of the final output gas from the endotracheal tube was about <NUM>, <NUM>, and <NUM>% in the O<NUM>-air-He mixture.

The feed gas of O<NUM> and air mixture or He was humidified by bubbling through DI water. The relative humidity (RH) of the humidified gas was measured constantly with a humidity sensor.

Thiazolyl blue tetrazolium bromide (MTT) was purchased from Sigma-Aldrich (St. Louis, MO, USA) and viability assays were carried out <NUM> hours after CAP treatment according to the manufacturer's protocol. Results were read by a BioTek microplate reader at <NUM> for absorbance.

The viability of A549 cells treated by the present invention were plotted, as shown in <FIG>. <FIG> shows the viability of A549 cells treated by the setup of the present invention with air mixture humidity adjustment for up to <NUM> minutes. The viability of A549 cells was decreased gradually with increasing treatment time. With <NUM> of treatment, the viability was reduced to <NUM>% (compared to no treatment). Oxygen fraction increasing in the gas mixture has indicated a weakened effect of the treatment from the <FIG>.

The same setup with <NUM>% O<NUM> in the feeding gas was then used to treat the cells for up to <NUM> (<FIG>). Cell viability was reduced below <NUM>% after <NUM> of treatment compared to no treatment, and the cells were completely eliminated after <NUM> of treatment.

When treated with the setup where only the helium is humidified, with helium humidity adjustment for <NUM>, all the cancer cells were eliminated by the CAP treatment as well (data shown in <FIG>).

A549 cells were treated with <NUM>% humidified Air/O<NUM> mixture (<NUM>:<NUM> v/v) at a flow rate of <NUM> LPM (O<NUM> fraction <NUM>%). Helium flow rate was <NUM> LPM and humidity was set at <NUM>%, <NUM>%, and <NUM>%.

A549 cells without treatment attached firmly to the culture dish, and nuclei were intact. The control in this experiment was Air/O<NUM> and He mixture-treated for <NUM>. Cells did not demonstrate any morphological changes compared to no treatment.

When He humidity was set at <NUM>% (dry helium), the cells started to shrink within <NUM> of cold plasma system treatment, but a significant amount of the cells were still viable. After increasing the treatment time to <NUM>, cell death was identified.

When He humidity was increased to <NUM>%, at <NUM> of treatment time, the cells demonstrated shrinkage and blebbing of the membrane. Cell shrinkage was more severe at <NUM> treatment time, and dead cells were visualized in a floating pattern.

When He humidity was increased to <NUM>% at <NUM> or <NUM> of treatment, almost all the cells were fragmented and not viable.

An MTT viability assay was performed on the cells. The results are shown in <FIG>. He humidity at <NUM>% (dry helium) and <NUM> treatment time reduced the viability to <NUM>%, and at <NUM> the viability was reduced the viability to <NUM>% compare to no treatment. When He humidity was set to <NUM>% or <NUM>%, there were no viable cells at <NUM> or <NUM> of treatment.

A more comprehensive study was performed to determine the minimum treatment time required for elimination of A549 cells. A549 cells were treated for <NUM>-<NUM> with humidified or dry Air/O<NUM> mixture (<NUM>:<NUM> v/v) at a flow rate of <NUM> LPM (O<NUM> fraction <NUM>%). Helium flow rate was <NUM> LPM and humidity was set at <NUM>%, <NUM>%, and <NUM>%. Images were taken <NUM> hour-post CAP treatment.

Phase contrast images of the A549 cells treated with a system in accordance with a preferred embodiment of the present invention for <NUM> - <NUM> with humidified or dry Air/O<NUM> mixture and various humidity of He were taken. When He humidity was set at <NUM>% (dry helium), cell number started to decrease at <NUM> of treatment, but cell morphology did not change significantly; when He humidity was set at <NUM>%, cell number started to decrease at <NUM> of treatment; when He humidity was set at <NUM>%, cell number started to decrease at <NUM> of treatment, and cell membrane and nuclei started to shrink significantly at <NUM> of treatment. Humidity of Air/O<NUM> did not induce significant morphological changes.

An MTT viability assay was performed on the cells (<FIG>). He humidity at <NUM>% (dry helium) did not induce much cell death compared to no treatment even at <NUM> of treatment. When He humidity was set to <NUM>%, cell viability gradually decreased with increasing of treatment time. About <NUM>% of cells were viable at <NUM> of treatment. When He humidity was set to <NUM>%, <NUM> of CAP treatment was able to reduce viability to below <NUM>%, and <NUM> of treatment completely eliminated the cells. Humidity of Air/O<NUM> did not result in significant differences in viability data. Based on these results, one may conclude that humidification of helium is a critical factor for the cold plasma system to eradicate lung cancer cells.

A cold atmospheric plasma system for treatment of respiratory infections in accordance with a first preferred embodiment of the present invention is described with reference to <FIG>. A helium gas source <NUM> is split into two lines <NUM>, <NUM>, with each of the two lines controlled by a mass flow controller (MFC) <NUM>. The Helium gas flow (<NUM> to <NUM>/min) in line <NUM> is passed through an H<NUM>O filled container (Humidifier 1130a) and then fed into a mixing chamber 1140a. The helium gas flow in the line <NUM> is fed directly into the mixing chamber 1140a. In this manner, with the mass flow controllers <NUM> on the lines <NUM>, <NUM> a relative H<NUM>O saturation in the gas exiting the chamber 1140a can be adjusted. Adjustment of the gas flow in the two lines <NUM>, <NUM> makes the overall flow rate and humidity fine tuning of the gas flow exiting the chamber 1140a possible. The humidity may be in the range of <NUM>%-<NUM>% with a preferred humidity of at least <NUM>%. The total helium flow in this embodiment could be varied from <NUM>/min to <NUM>/min in all cases. The humidity of Helium gas in the chamber 1140a may be measured, for example, via calibrated High-Accuracy Humidity and Temperature Meter (not shown). The humidified helium gas from the chamber 1140a is fed into an electrosurgical generator <NUM>, referred to herein as a "Cold Atmospheric Plasma (CAP) Generator. " A variety of electrosurgical generators are known in the art and could be used with the present invention. The gas being fed into the Cold Atmospheric Plasma (CAP) Generator <NUM> is referred to herein as the "carrier gas.

At the same time, an un-humidified air supply <NUM> (feed gas) is controlled by a mass flow controller (MFC) <NUM>. The air gas flow is passed through a second H<NUM>O filled container (Humidifier 1130b) and then is fed into a mixing chamber 1140b. Also at the same time, a source <NUM> of an unhumidified pressurized third gas, oxygen in this case, is connected to a third H<NUM>O filled container (Humidifier 1130c). The humidified third gas (oxygen) is fed into chamber 1140b where it mixes with the humidified air. In this manner, with the mass flow controllers <NUM> on the air and oxygen lines the relative oxygen percentage exiting the chamber 1140b can be adjusted. The humidity of each of the air and oxygen may be in the range of <NUM>%-<NUM>% with a preferred humidity of at least <NUM>%. The humidity of mixture in the chamber 1140b may be measured, for example, via a calibrated High-Accuracy Humidity and Temperature Meter (not shown) and the oxygen content may be measured for example with an oxygen sensor. The humidified air and oxygen from the chamber 1140b are provided to a respiratory delivery system <NUM>, such as a ventilator, CPAP machine, BIPAP machine, or other known respiratory deliver system. The respiratory delivery system <NUM> will mix, adjust and measure the pressure, flowrate, ratio and frequency of the patient inbreath of exhaust air, oxygen and CO2. The output of the respiratory delivery system <NUM> is connected to the CAP joint mixer, for example, via tubing <NUM> and connector <NUM>.

Ozone (O<NUM>) generated by a system in accordance with a preferred embodiment of the present invention was measured at the end of the endotracheal tube with an ozone detector (Forensics Detectors, CA). The measurement was carried out with all settings tested above, i.e., CAP was set to <NUM> V with humidified or dry Air/O<NUM> mixture (<NUM>:<NUM> v/v) at a flow rate of <NUM> LPM (O<NUM> fraction <NUM>%) and helium flow rate was <NUM> LPM and humidity was set at <NUM>%, <NUM>%, and <NUM>%. Ozone level was shown in <FIG>. At the same helium humidity, dry Air/O<NUM> yielded higher O<NUM> level compared to humidified Air/O<NUM>. Higher humidity of helium generated higher concentration of O<NUM>, which resulted in stronger reduction effect on the cells as shown earlier in <FIG>. This correspondence indicates that O<NUM> is a critical species in the cocktail that generated by a system in accordance with a preferred embodiment of the present invention. <FIG> shows ozone production rate significantly decreased at lower voltage. Therefore, <NUM> or <NUM> V was used to test cell viability instead of <NUM> V for safety purpose (<FIG> and <FIG>). O<NUM> production rate was higher when Air and O<NUM> were fed into the system as a mixture (<FIG>) compared to separate Air and O<NUM> infusion (<FIG>).

However, the cold plasma system with previously demonstrated settings, i.e., CAP was set to <NUM> V with humidified or dry Air/O<NUM> mixture (<NUM>:<NUM> v/v) at a flow rate of <NUM> LPM (O<NUM> fraction <NUM>%) and helium flow rate was <NUM> LPM and humidity was set at <NUM>%, <NUM>%, and <NUM>%, produced a large amount of ozone (data demonstrated in Section <NUM>) which is over the safety limit by OSHA standard. In order to lower the ozone generation, lower voltage (<NUM> - <NUM> V) was utilized to treat the cells. Because the presence of oxygen in the Air/O<NUM> mixture inflamed the ozone production, Air and O<NUM> were fused into the joint CAP mixer separately to lower the O<NUM> formation. The viability data of A549 cells treated at settings with low O<NUM> level (i.e. lower voltage and separation of Air and O<NUM>) is shown in <FIG>.

When Air and O<NUM> were added to the system as a mixture (<FIG>), the capability of CAP on reducing cancer viability was higher compared to where Air and O<NUM> were fused to the system separately (<FIG>). CAP treatment of <NUM> at <NUM> V with Air/O<NUM> mixture or <NUM> at <NUM> V with Air and O<NUM> separation were able to lower cancer cell viability to less than <NUM>% percent.

A system in accordance with a preferred embodiment of the present invention was used to treat <NUM> Phosphate Buffer Saline (PBS) in <NUM>-well plates with Argon Coagulation Mode for <NUM> or <NUM> continuously and in intervals. For interval treatment, CAP was administered in <NUM> + <NUM> + <NUM> or <NUM> + <NUM> + <NUM> + <NUM> manner with <NUM> break between each interval. Voltage was set at <NUM> or <NUM> V. Helium, O<NUM> and air were all humidified. Flow rate of helium was set at <NUM> LPM. Oxygen and air flow rate were both set at <NUM> LPM.

Among the cocktail plasma-generated reactive oxygen species (ROS) and nitrogen species (RNS) in the treated solution, hydrogen peroxide (H<NUM>O<NUM>), nitrite (NO<NUM>-) and nitrate (NO<NUM>-) are the most commonly studied long-lived species. Their concentrations were measured in treated PBS using colorimetric Hydrogen Peroxide Assay Kit (Sigma-Aldrich, MAK311-1KT), Griess Reagent System (Promega G2930) and colorimetric Nitrite/Nitrate Assay Kit (Sigma-Aldrich <NUM>), with CAP on or off. Results were read by a BioTek microplate reader at <NUM>, <NUM>, and <NUM>/<NUM> for absorbance, respectively.

The concentrations of H<NUM>O<NUM>, NO<NUM>- and NO<NUM>-with a system in accordance with a preferred embodiment of the present invention were plotted. Previous viability data demonstrated that at <NUM> V, continuous treatment for <NUM> with Air and O<NUM> mixture setup or <NUM> with Air and O<NUM> separation setup can both lower the viability of A549 to less than <NUM>%. As indicated in <FIG>, <NUM> of Air and O<NUM> mixture setup at <NUM> V generated <NUM> H<NUM>O<NUM>, <NUM> NO<NUM>- and <NUM> NO<NUM>- compared to <NUM> + <NUM> + <NUM> interval treatment generated <NUM> H<NUM>O<NUM>, <NUM> NO<NUM>- and <NUM> NO<NUM>-, whereas continuous <NUM> of Air and O<NUM> separation setup at <NUM> V generated <NUM> H<NUM>O<NUM>, <NUM> NO<NUM>- and <NUM> NO<NUM>- compared to <NUM> + <NUM> + <NUM> + <NUM> interval treatment generated <NUM> H<NUM>O<NUM>, <NUM> NO<NUM>- and <NUM> NO<NUM>-. Nitrate (NO<NUM>-) were too low to detect in most of the settings. Gas mixture alone does not generate significant amount of ROS or RNS.

In the case of <NUM> continuous treatment, H<NUM>O<NUM> was generated in <NUM> of media by CAP treatment with <NUM> LPM of gas flow. The detected species were as follows:.

In all cases, <NUM>, <NUM>, <NUM>, and <NUM>/m<NUM> are lower than NIOSH and OSHA permissible exposure limit for H<NUM>O<NUM>, which is <NUM>/m<NUM> (https://www. gov/niosh/npg/npgd0335.

Claim 1:
A system for performing plasma treatment of respiratory infections comprising: an electrical energy generator (<NUM>) configured to generate electrical energy to plasmatize a carrier gas into a plasma; and a dielectric barrier discharge mixer (<NUM>) comprising: an interior chamber formed from a dielectric, said interior chamber having a first input (<NUM>) configured to fluidly connect to a source of a humidified carrier gas (<NUM>), a second input (<NUM>) configured to connect to a source of a humidified feed gas (<NUM>), and an output (<NUM>) configured to connect to a delivery member (<NUM>); an active electrode (<NUM>) inside said interior chamber and connected to an electrical output of said electrical energy generator; and an outer electrode (<NUM>) connected to a ground; wherein a plasma is generated in said interior chamber when electrical energy is supplied from said electrical energy generator (<NUM>) to said interior electrode (<NUM>) while both humidified feed gas and humidified carrier gas flow into said interior chamber, and further comprising: a first humidifier (<NUM>) fluidly connected to said first input (<NUM>) of said chamber in said dielectric barrier discharge assembly (<NUM>); and a second humidifier (<NUM>) fluidly connected to said second input (<NUM>) of said chamber in said dielectric barrier discharge assembly (<NUM>).