MEASUREMENT OF CARBON DIOXIDE IN BLOOD

Abstract: A method of determining the total carbon dioxide concentration in plasma (TCO2 plasma) directly from a whole blood sample involves adjusting TCO2 measured in the whole blood sample (TCO2 whole blood) using a volume dilution factor (VDF) to give a value that is equivalent to TCO2 plasma.

Description: MEASUREMENT OF CARBON DIOXIDE IN BLOOD The invention relates to the determination of the total carbon dioxide concentration in a whole blood sample. Background of the Invention The measurement of total carbon dioxide concentration (TC0 2 ) in serum or plasma is a routine analysis performed in clinical laboratories as a patient profile test. In particular, measurement of TC0 is useful in determining the acid-base status of a patient. For example, plasma TC0 2 can be used by a physician in the diagnosis of a patient in renal failure or acute acidosis. TC0 2 in serum or plasma is comprised of three major chemical forms: dissolved C0 2 (3%) ; carbamino (R- NHCOO " ) derivatives of plasma protein (33%) ; and bicarbonate (HC0 3 " ) anion (64%) . Other minor forms include carbonic acid (H 2 C0 3 ) and carbonate anion (C0 3 = ) . Most of the procedures that quantify TC0 2 in serum or plasma involve acidification of the sample to convert all of the C0 2 -containing species to C0 2 gas, as is summarized in equation (1) : C0 2 + R-NHCOO " + HC0 3 " + H-.CO 3 + C0 3 = + Acid → C0 2 t + H 2 0 (1) Once liberated, the C0 2 gas can be measured manometrically in batch analysis or by continuous-flow procedures where the C0 gas diffuses across a membrane into a bicarbonate-containing solution thereby lowering the pH of the solution. The change in pH can be detected either potentiometrically using a pH glass electrode or spectrophotometrically if the bicarbonate buffer contains a pH indicating dye. Additionally, there are detection methods that use enzymes or bicarbonate-selective electrodes to quantify TC0 2 . These techniques apply to serum or plasma samples. Therefore, centrifugation of the whole blood sample to separate the cells and plasma prior to analysis is required. Summary of the Invention The invention relates to the determination of TC0 2 in plasma or serum directly from a whole blood sample. The TC0 2 in plasma is essentially equivalent to the TC0 2 in serum; TCO- plasma will be used herein to denote either value. The method involves adjusting TC0 2 measured in the whole blood sample (TC0 2 wh oie bl oo d ) us;Ln 9 a volume dilution factor (VDF) to provide TC0 2 plasma . The VDF is determined for each whole blood sample and, preferably, can be related directly to the hematocrit (Het) of that sample. An advantage of the present invention is that TC0 2 piasrrιa can be determined accurately and directly from a whole blood sample. Since the measurement does not depend on plasma or serum samples, the method of the invention avoids time-consuming sample preparation steps. Other features and advantages of the invention will be apparent from the description of the preferred embodiment thereof, and from the claims. Brief Description of the Drawing The Figure is a fluid flow diagram. Description of the Preferred Embodiments The preferred method includes measuring τco 2 w h o l e bl oo d ^ n a wnole blood sample, generating a VDF from the Het of the same whole blood sample, and adjusting the TC0 2 whole bl oo d us i nc J tne VDF to provide the TC0 2 plaB m of tne whole blood sample. Methods for measuring TC0 2 are described in U.S. Pat. Nos. 3,973,912, 4,321,545 and 5,429,930, each of which is incorporated by reference. The preferred method of measuring TC0 2 whole blood involves acidification of the whole blood sample to liberate C0 2 gas from the sample according to equation (1) . The sample and acidifying agent mix as they pass through a mixing chamber. Acidifying agents include aqueous solutions of sulfuric acid, citric acid, lactic acid and combinations thereof. The preferred acidifying agent is an aqueous solution containing 90 mM citric acid, although the concentration can be varied over a wide range. The preferred TC0 2 gas electrode includes a pH glass electrode along with a Ag/AgCl reference electrode. The two electrodes are covered by a C0 2 gas permeable membrane such as silicone membrane. A bicarbonate/chloride internal filling solution (IFS) is trapped between the membrane and the pH electrode. The C0 2 gas generated by acidification passes through the silicone membrane. It equilibrates with the IFS. The pH of the IFS will change with the C0 2 concentration. By measuring pH with the pH glass electrode, the C0 2 concentration can be determined. The TC0 2 gas electrode is calibrated before use. Calibration involves using two standard aqueous bicarbonate solutions. Each standard solution, when mixed with acid, will produce a known C0 2 concentration which in turn will produce a known pH in the IFS trapped between the membrane and the pH electrode. Once the TC0 2 electrode is calibrated, TC0 2 who i e blood ^ s determined comparing the unknown with one calibration standard. TC0 2 plasma ^ s obtained by adjusting the measured TC0 2 whole blood us;Ln< -J tne VDF of tne same sample. The preferred adjustment is shown in equation (2) : TC0 2 plasma - TC0 2 whole blood / VDF (2) The VDF represents the fractional volume of the extracellular fluid in the whole blood sample. Preferably, the VDF is determined from the Het of the same whole blood sample according to equation (3) : VDF = 1 - Het / 100 (3) Het is the volume of red blood cells occupying the total volume of the whole blood sample reported as a percentage. Het measurement is performed routinely in clinical laboratories using various methods to diagnose conditions such as anemia or polycythemia. The preferred method of determining Het is by measuring the conductivity of the whole blood sample using a pair of calibrated electrodes as disclosed in U.S. Pat. No. 4,686,479. The Het sensor may be factory calibrated or calibrated using aqueous standards. TC0 2 pi a g ma obtained from analyzing a whole blood sample following the method of the invention is equivalent to the TC0 2 determined after isolating the plasma from the whole blood sample by centrifugation. Table I lists TC0 2 data of patients that was measured in whole blood and in the separated plasma. The plasma samples acquired from the corresponding blood samples were subsequently analyzed under identical conditions to obtain the directly measured TC0 2 p asma ' Hct °^ ^ ne wn °l e blood sample was measured by the method disclosed in U.S. Patent No. 4,686,479. Finally, the TC0 2 who e bl oo d was adjusted according to equations (2) and (3) . The data in Table I indicate that TC0 2 pl as a measured using the preferred method correlates with TC0 2 pl asm measured directly in plasma samples. The method of the invention can be used in most typical blood chemistry analyzers. The features of a preferred blood chemistry analyzer are presented in the figure, which shows the preferred arrangement of pumps and analysis chambers in the analyzer. The analyzer includes two peristaltic pumps and a series of analysis chambers. Pump 1 draws the sample through tube 10 into analysis chamber 12 where the Het of the sample is determined if the sample is a whole blood sample. The sample then flows through a set of analysis chambers 14 where other sample chemistry is probed. The sample and acidifying agent are then combined in mixing chamber 16. The acidifying agent is delivered to chamber 16 by pump 2. The mixture flows to chamber 18 which houses a TC0 2 gas electrode. The liberated C0 2 gas is measured in chamber 18 using a TC0 2 gas electrode, and related to TC0 2 in the sample. The TC0 2 in the whole blood sample is then converted to TC0 2 p i aSma us i n 9 tne data collected for that sample. The TC0 2 p asma measurement described above can be incorporated into a NOVA CRT12 or CRT14 Chemistry Analyzer, which is available from Nova Biomedical Corp., with appropriate modification. The analyzer allows most of the blood chemistry to be determined in a single run of a whole blood sample, decreasing sample analysis time which is critical in emergency situations. Other embodiments are within the claims. What is claimed is:

Claims: 1. A method of determining the total C0 2 concentration in plasma of a whole blood sample, said whole blood sample containing red blood cells and plasma, wherein the fractional volume of said plasma in said whole blood sample comprises the volume dilution factor of said whole blood sample, said method comprising: (a) loading an uncentrifuged whole blood sample containing red blood cells into a blood chemistry analyzer including a C0 2 sensor and a hematocrit- measuring station; (b) measuring the total C0 2 concentration of said uncentrifuged whole blood sample containing red blood cells with said C0 2 sensor; (c) measuring the hematocrit of said uncentrifuged whole blood sample containing red blood cells at said hematocrit-measuring station; (d) generating the volume dilution factor of said uncentrifuged whole blood sample containing red blood cells from said hematocrit; and (e) adjusting the C0 2 concentration of said uncentrifuged whole blood sample containing red blood cells using said volume dilution factor to provide the total C0 2 concentration in plasma of said uncentrifuged whole blood sample containing red blood cells. 2. The method of claim 1, wherein the total C0 2 concentration of the whole blood sample is determined by the addition of an acidifying agent to said whole blood sample to liberate C0 2 gas which is quantified by a detector. 3. The method of claim 2, wherein the acidifying agent is an aqueous solution comprised of an acid more acidic than carbonic acid. 4. The method of claim 3, wherein the acid is citric acid. 5. The method of claim 4, wherein the acidifying agent is an aqueous solution containing sulfuric acid. 6. The method of claim 3, wherein the detector of liberated C0 2 gas is a manometer. 7. The method of claim 3, wherein the detector of liberated C0 2 gas is a pH glass electrode. 8. The method of claim 2, wherein hematocrit is determined by impedance or conductance.

Inventors: CHIEN JEFFREY, YOUNG CHUNG CHANG, CHIEN, JEFFREY, YOUNG, CHUNG, CHANG
Applicants: NOVA BIOMEDICAL CORP, NOVA BIOMEDICAL CORPORATION
Application year: 1996
Main classification: G01N33
IPC codes: G01N33