Abstract:
A device having a catheter and a hub which facilitates the transfer of an embryo containing solution into the uterus of a mammal, for example, a small mammal such as a rat or mouse. The catheter&#39;s shape and tip is designed to simplify insertion and minimize injury to the mammal. The hub is designed to further control the insertion of the catheter and to interface with a pipette, for example, that provides the force required for the transfer of an embryo containing solution. As a result, the transfer can occur without requiring surgery.

Description:
RELATED APPLICATIONS 
     This application claims priority to U.S. Application Ser. No. 61/128,726 filed May 23, 2008 the disclosure of which is incorporated herein by reference in its entirety. 
    
    
     BACKGROUND OF THE INVENTION 
     Field of the Invention 
     The present invention relates generally to embryo transfer techniques and more particularly with a device and method for accomplishing such a transfer without surgery. 
     Description of Related Art 
     Existing methods in rodent reproductive sciences have a number of shortcomings that make them expensive, difficult, and inefficient. It generally takes about 30 minutes to transfer embryos to female mice using surgical procedures, although this will vary depending on the expertise of the person performing the technique. Also, surgical embryo transfer involves anesthesia, post-operative care, the use of analgesics, and possible complications including infection. The significance of these issues continues to grow as regulations regarding the use and care of experimental animals expand. Anesthetics and analgesics are relatively expensive as well and some are considered controlled substances, so the elimination of these reagents represents a considerable cost savings and reduction of regulations. 
     Furthermore, the technical expertise required for surgical transfer of embryos is considerably significant and, thus, limits the number of potential users of genetically modified mice. 
     Thus, there remains the need for devices and methodology that eliminates the need for anesthesia, surgery, and the use of analgesics, and dramatically reduces the time needed for post-operative care and essentially eliminates infection and other post-operative complications. Also a reduction in the technical expertise required for surgical transfer of embryos would help expand the number of potential users of genetically modified mice and other animals. 
     BRIEF SUMMARY OF THE INVENTION 
     Embodiments of the present invention relate to a device, and a method for its use, having a catheter and a hub which facilitates the transfer of an embryo containing solution into the uterus of a mammal, for example, a small mammal such as a rat or mouse. The catheter&#39;s shape and tip is designed to simplify insertion and minimize injury to the mammal. The hub is designed to further control the insertion of the catheter and to interface with a pipeter, for example, that provides the force required for the transfer of an embryo containing solution. As a result, the transfer can occur without requiring surgery. 
     It is understood that other embodiments of the present invention will become readily apparent to those skilled in the art from the following detailed description, wherein it is shown and described only various embodiments of the invention by way of illustration. As will be realized, the invention is capable of other and different embodiments and its several details are capable of modification in various other respects, all without departing from the spirit and scope of the present invention. Accordingly, the drawings and detailed description are to be regarded as illustrative in nature and not as restrictive. 
    
    
     
       BRIEF DESCRIPTION OF DRAWINGS 
       Various aspects of a non surgical embryo transfer device are illustrated by way of example, and not by way of limitation, in the accompanying drawings, wherein: 
         FIG. 1  depicts a non-surgical embryo transfer device in accordance with the principles of the present invention. 
         FIG. 2  depicts the device of  FIG. 1  in addition to a speculum. 
         FIG. 3  depicts the device of  FIG. 1  in relation to insertion in an animal in accordance with the principles of the present invention. 
         FIG. 4  depicts another embodiment of a non-surgical embryo transfer device in accordance with the principles of the present invention. 
     
    
    
     DETAILED DESCRIPTION OF INVENTION 
     The detailed description set forth below in connection with the appended drawings is intended as a description of various embodiments of the invention and is not intended to represent the only embodiments in which the invention may be practiced. The detailed description includes specific details for the purpose of providing a thorough understanding of the invention. However, it will be apparent to those skilled in the art that the invention may be practiced without these specific details. In some instances, well known structures and components are shown in block diagram form in order to avoid obscuring the concepts of the invention. 
     Embodiments of the present invention relate to intrauterine transfer of embryos in animals, for example in animals such as small rodents, including but not limited to mice and rats. Other animals in addition to rodents are contemplated within the scope of the present invention as well. 
     A small, flexible plastic device has been developed to physically introduce a catheter through the cervix of the small rodent, which can then be used to transfer a solution containing embryos into the uterus. Once transferred, the embryos attach to the endometrial tissue, implant and develop. 
     This device has a number of beneficial characteristics that make it well-suited to traverse the cervix without damaging the cervical or uterine tissue. For example, the length of the device is designed, such that it is long enough to get past the cervix but not too long to damage endometrial tissue, this allows the tip to be located within the uterus at a position which improves the likelihood of successful embryo transfer. Also, in at least one embodiment, the hub interacts with a separate speculum to further assist with correctly positioning the device in a female rodent, or animal, for embryo transfer. Additionally while a non-flexible catheter may be used without departing from the scope of the present invention, the use of flexible tubing beneficially provides for easier insertion and a reduction in potential tissue damage. Because there are currently apparatus available that can provide the correct amounts of an embryo solution, the hub of the device may be configured to attach to such apparatus. Alternatively, the catheter and the apparatus providing the embryo solution may be configured as an integral unit. 
     In practice, the device can be inserted through the cervix where embryos can then be expunged. Afterwards, the device can be removed. Accordingly, one of ordinary skill will appreciate that all three steps can be performed in non-anesthetized female animals. The lack of anesthesia provides a number of benefits over current surgical techniques for embryo transfers. These existing methodologies for embryo transfer in small rodents require the surgical transfer into anesthetized females. Anesthetics are considered to be “controlled substances” under DEA guidelines, which require specialized handling and storage. 
     As mentioned, embryo transfer using embodiments of the present invention eliminates the need not only for anesthesia but also for surgery. All current methods of embryo transfer in small rodents require surgical transfer. The surgical methods in small rodents are technically challenging and require substantial training and a high level of skill. Surgical transfer requires that females be anesthetized, as described above, and complications (mistakes during surgery, inappropriate levels of anesthesia, post operative infection) can lead to death of female mice. Surgery also leads to post-operative care of female mice. This care requires monitoring of the mice at regular intervals for several weeks post-surgery to insure that they are healthy and not suffering any complications (including infections) and the possible use of analgesics. Animal use guidelines also require that surgical instruments are sterile prior to use; sterilization requires autoclaves, which are expensive and shorten the usable life of costly high-precision surgical instruments. 
     An additional benefit of practicing the present invention is that it results in a dramatic time savings; surgical transfer require about 30 to 45 minutes of labor for each animal whereas use of embodiments of the present invention allows the transfer to be accomplished in less than one minute. 
     The use of genetically modified mice is well established as a critical aspect of biomedical research, and the use of such mice is expected to rise. At the same time, animal research in academic and private institutions is strictly regulated to insure proper treatment of animals, and guidelines for animal use continue to increase. Academic institutions are required to have IACUC (Institutional Animal Care and Use Committees) oversight to insure proper and humane use of animals. While it is acknowledged that animal research is an essential component of biomedical research, the use of animals is guided by “the three R&#39;s”. These include replacement (using non-animal alternatives), reduction (the use of fewer animals to produce the desired result) and refinement (to reduce the incidence or severity of inhumane or painful procedures). Embodiments of the present invention completely eliminate the need for surgery while performing embryo transfer and provide an alternative, non-surgical transfer that is a much more humane, much less painful method. 
       FIG. 1  depicts a device  100  configured in accordance with the principles of the present invention. There is a hub portion that has a first end  102  and a second end  104 . Extending from the second end  104  is a catheter  106  the terminates at a distal tip  108 . The length  112  of the catheter  106  is discussed in detail below. The hub portion includes a cavity  110  that is in fluid communication with the inside passageway of the catheter  106 . In this way, fluid or other material may be drawn into, and expelled from, the catheter tip  108  by application of the appropriate pressure at the cavity  110 . 
     The first end  102  of the hub portion may be configured to be coupled with a pipeter or similar device that can be used to supply material into the cavity  110 . The design of the first end  102  of the hub portion is not as critical; it is designed so that the hub portion may fit snugly on a pipeter such as, for example, a Gilson PIPETMAN® pipeter. However, other pipeters are available, and designing the first end  102  for use with other pipeters is contemplated within the scope of the present invention. 
     In addition to a pipeter, one of ordinary skill will recognize that there are a number of functionally equivalent devices for delivering an embryo solution that can be used instead of the pipeter without departing from the scope of the present invention. In yet another alternative shown in  FIG. 4 , a separate pipeter, or similar, device is not used. Rather, a single device  400 , having an integrated pipeter  402  and transfer device  404 , is used to carry out the entire procedure of loading and dispensing the embryo-containing solution, negating the need for a separate pipeter. 
     It is beneficial that the tip  108  is tapered in order to get through the cervix. The degree of the taper may be modified for devices depending on the type of animals on which they are used. As for manufacturing and material, the catheter  106  and tip  108  may be extruded from fluorinated ethylene propylene or FEP (TEFLON®) while the hub portion may be made from polyethylene, or the like. This selection of material advantageously provides a tip  108  that has rigidity but also some flexibility. 
     In  FIG. 2 , the catheter  106  is shown in relation to a speculum  202 . In operation, the tip  108  of the catheter  106  extends into a proximate end  206  of the speculum  202  and exits out a distal end  204 . In relation to being inserted in an animal, the distal end  204  may also be called the “front” end of the speculum  202 . The speculum, which is inserted in the vagina of the animal, can be constructed of any semi-rigid plastic or composite material suitable for the intended environment. It can be constructed so as to be a disposable item or, alternatively, have a robust enough construction to allow sterilization and re-use. Furthermore, the speculum can be initially configured to be attached to the device prior to use or it can be, as mentioned above, a separate piece. 
     As shown, the device has a passageway within the hub and catheter that allows a solution received at the tip of the catheter to be drawn up into the catheter. In this way, the pipette and the device can deliver a embryo solution within an animal&#39;s uterus when the device is inserted through the cervix of that animal. 
     One beneficial aspect of the device  100  of  FIG. 3  is the distance  310  from the end of the tip  108  to the front end  204  of the speculum  202 . The device is sized such that when the speculum  202  is placed in the mammal&#39;s vagina  302 , the distance  310  is sufficiently long to get past the cervix  304  and transfer the embryos into the uterus  306 , but not so long as to extend far enough to damage the uterine wall  308 . Thus, the back end  206  of the speculum  202  acts as a positive stop preventing the tip  108  from extending so far into the uterus as to cause damage. The flexibility of the tip  108  also helps prevent damage even if the uterine wall  308  is contacted. 
     One example mammal that the device can be used with is CD-1 mice. Because of slight physiological differences between different strains of mice, the distance  310  of the device  100  may vary slightly for different mouse strains. The distance  310  can be determined by performing testing on euthanatized female mice of different strains. In addition to CD-1 mice, the device can also be used for transcervical transfer of embryos in rats, other types of mice, and other mammals. Accordingly, because of the size difference between mice, rats and other mammals, a device  100  that is appropriate for each such animal will have its own, respective distance  310  that will differ among such devices. 
     By way of example, for CD-1 mice, the device  100  of  FIG. 1  may have the following dimensions. One of ordinary skill will recognize that such dimensions are provided as the best mode known for use with a CD-1 mouse; however, embodiments of the present invention are not limited to only these specific dimensions. As discussed herein, the device  100  is sized to accommodate different species of animals in such a way as to allow embryos to be transferred past the cervix but without causing damage to the uterine walls. The overall length of the device  100  may be between about 1.6 and 1.7 inches with the hub portion  102 ,  104  being about 0.625 inches in length. Thus, the distance  112  of the catheter portion  106  may be about 0.9 to 1.1 inches. As for diameter, the catheter  106  may have an inner diameter (ID) of about 0.018 to about 0.021 inches and an outer diameter (OD) of about 0.028 to about 0.031 inches. As shown, the tip  108  tapers towards its end and this tapered section may be about the last 0.05 inches of the tip  108 . As mentioned, these specific dimensions may be varied for various animals, and for mice as well, without departing from the scope of the present invention. 
     Also, one other factor to consider when sizing the device  100  for different animals is that the size of the speculum will likely be different for different animals. By way of example, with CD-1 mice, there are two differently sized speculums that may be used with the device. A smaller speculum is used to help open the vagina of the mouse. The smaller speculum dimensions may be about 0.087 inches ID, 0.109 inches OD, and a length of about 0.475 inches. As discussed, the device  100  and the speculum are designed to assist in the correct depth placement of the catheter inside the uterus. In one example, the larger speculum is sized appropriately with dimensions of about 0.140 inches ID, about 0.162 inches OD, and a length of about 0.400 inches. As for material, the speculums can be made of polyethylene or polypropylene. 
     As discussed above, one particular use of the device  100  is for non-surgical transfer of embryos into female mice. However, other embodiments of the invention contemplate that the device may be used for the transfer of sperm (fresh and cryopreserved) or for the transfer of cryopreserved blastocysts as well as embryos that are obtained by in vitro fertilization. 
     In at least one embodiment, the device is packaged in a sterile packaging and is intended to be disposed after each use. The user of the device may benefit from using a magnifying device when drawing the embryos into the catheter, but such an accessory is not required to practice the present invention. 
     The previous description is provided to enable any person skilled in the art to practice the various embodiments described herein. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments. Thus, the claims are not intended to be limited to the embodiments shown herein, but are to be accorded the full scope consistent with each claim&#39;s language, wherein reference to an element in the singular is not intended to mean “one and only one” unless specifically so stated, but rather “one or more.” All structural and functional equivalents to the elements of the various embodiments described throughout this disclosure that are known or later come to be known to those of ordinary skill in the art are expressly incorporated herein by reference and are intended to be encompassed by the claims. Moreover, nothing disclosed herein is intended to be dedicated to the public regardless of whether such disclosure is explicitly recited in the claims. No claim element is to be construed under the provisions of 35 U.S.C. §112, sixth paragraph, unless the element is expressly recited using the phrase “means for” or, in the case of a method claim, the element is recited using the phrase “step for.”