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Publication numberUS3629142 A
Publication typeGrant
Publication dateDec 21, 1971
Filing dateDec 8, 1969
Priority dateDec 8, 1969
Publication numberUS 3629142 A, US 3629142A, US-A-3629142, US3629142 A, US3629142A
InventorsMarbach Edward P
Original AssigneeMarbach Edward P
Export CitationBiBTeX, EndNote, RefMan
External Links: USPTO, USPTO Assignment, Espacenet
Reference standard blood serum for the calibration of automatic blood serum analyzing apparatus
US 3629142 A
Abstract  available in
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Claims  available in
Description  (OCR text may contain errors)

United States Patent 3,629,142 REFERENCE STANDARD BLOOD SERUM FOR THE CALIBRATION OF AUTOMATIC BLOOD SERUM ANALYZING APPARATUS Edward P. Marbach, 4607 Marwood Drive, Los Angeles, Calif. 90065 No Drawing. Filed Dec. 8, 1969, Ser. No. 883,286 Int. Cl. G01n 31/22, 33/16 US. Cl. 252-408 5 Claims ABSTRACT OF THE DISCLOSURE A freeze-dried blood serum is provided for use as a reference standard in the calibration of automatic blood serum analyzing apparatus, comprising a human serum in which the levels of concentration of its constituents have been precisely adjusted, and which is specially processed for use in apparatus for the simultaneous analysis of the constituents of each of a succession of blood serum samples. The reference standard blood serum of the invention contains trihydroxymethylamine (Tris) carbonate so as to permit it to be reconstituted just prior to use with distilled water so as to restore a predetermined carbon dioxide level in the reference serum. Alternately, the reference standard blood serum contains Tris, and is reconstituted with distilled water and carbon dioxide.

BACKGROUND OF THE INVENTION Various types of apparatus have been developed in recent years for the automatic analysis of human blood serum, and which are capable of making rapid simultaneous analytical determinations on a precise quantitative basis of various components of each of a series of blood serum samples successively introduced into the apparatus. Biochemical determinations may be made, for example, by such apparatus, of the following constituents of the blood serum samples: albumin, alkaline phosphatase, bilirubin, calcium, chloride, cholesterol, carbon dioxide content, creatinine, glucose, lactic dehydrogenase (LDH), inorganic phosphorus, potassium, sodium, total protein, transaminase (SGOT), urea nitrogen, uric acid, creatine phosphokinase (CPK).

The aforesaid apparatus may be of the type described, for example, in the Skeggs et a1. Pat. 3,241,432 which issued Mar. 22, 1966, and which is assigned to the Technicon Instruments Corporation of Chauncey, N.Y. The Skeggs apparatus, for example, is capable of making simultaneously a multiplicity of different analyses of a blood serum sample, with respect to the constituents referred to, for example, in the preceding paragraph, and of rapidly repeating the analyses on successive samples.

In the operation of the apparatus described in the Skeggs patent, for example, a series of individual separate blood serum samples are successively introduced into the machine, and are formed into a stream which subsequently is divided into a plurality of separate streams, depending upon the number of separate tests to be made. Each of the streams is separately treated for quantitative analysis with respect to the particular constituents, and the results of the analyses are recorded, for example, on a chart of a recorder, or are otherwise stored in any prescribed correlated form.

As is well known, the determination of the carbon dioxide content of the blood is of great importance in many clinical procedures. The aforesaid apparatus is capable of determining the carbon dioxide content of the test sample. Therefore, it is important for the reference standard blood serum to include a predetermined carbon dioxide content so that it may be used to calibrate the apparatus for that particular determination. However, it

3,629,142 Patented Dec. 21, 1971 has been found that in the freeze-drying of the blood serum for reference standard purposes, a substantial amount of carbon dioxide content of the serum is lost in the form of gaseous CO In order to calibrate apparatus of the type described in the aforesaid patent, a reference standard blood serum sample is used whose constituents have precisely predetermined values. It is necessary for such reference standard serums to be capable of being stored for long periods of time without deterioration, and for that reason it has been the practice to freeze-dry them. It has also been the prior art practice to reconstitute the reference standard serum before use by means, for example, of a separate aqueous solution of ammonium bicarbonate, to restore the carbon dioxide level to a predetermined value.

However, a problem which has arisen when the abovedescribed prior art reference standard serum is used is that a separate vial of a measured amount of the ammonium bicarbonate solution had to be provided, and reliance had to be placed on the ultimate user that the reconstituting solution would be added in the proper quantity to the reference standard serum. This makes the reliability of the results of the apparatus dependent upon the ultimate user.

Moreover, when the prior reference standard serum is to be used for different analytical procedures, different levels of carbon dioxide contents are required, and this calls for different concentrations of the reconstituting solution. In such cases, different vials of different concentrations of the ammonium bicarbonate solution must be provided with the prior art reference standard blood serum, and there is no assurance that the ultimate user would use the proper vial of the reconstituting solution for the appropriate procedure.

The prior art provision of the separate aqueous solution of the ammonium bicarbonate for the reconstitution of the freeze-dried serum just prior to use is necessary, because the reconstituted prior art serum was found to be stable only for a few days, and then only if stored in refrigeration between, for example, 2 and 8 centigrade. It was found, for example, that storage of the reconstituted prior art reference serum at room temperature tended to alter the SGOT and LDH value in the reconstituted serum.

Also the ammonium bicarbonate could not be intermixed into the prior art reference serum originally in the laboratory because it is completely lost during freeze drying.

An important object of the present invention is to provide a reference standard serum to which a carbonate or bicarbonate of Tris(hydroxymethyl) aminomethane is added prior to when it is in its freeze-dried state, and which may be reconstituted merely by adding distilled water. The Tris carbonate has no effect on any of the constituents of the reference standard serum and does not affect the stability of its enzyme constituents in any way, even over long intervals of time. The use of Tris carbonate takes the concentration control of the reconstituting element out of the hands of the ultimate user, and into the laboratory where it can be established precisely and under optimum environmental conditions. The chemical name is 2-amino-2-(hydroxymethyl)-1,3-propandiol. The formula is It has been found that the use of the Tris carbonate reference standard serum of the present invention results in a reconstituted serum by the addition of distilled water, which functions with all the efficacy of the properly re constituted prior art ammonium bicarbonate reference standard serum. The reference standard serum of the present invention has no tendency to plug the apparatus in which it is used, and it is clear and colorless. so that it does not affect any of the colorimeter or other tests carried out by the apparatus. The use of Tris carbonate in the reference standard serum of the invention results in a proper pH, so that the stability of enzymes such as lactic dehydrogenase and transaminase, is in no way affected.

An important feature of the present invention is that the addition of a measured amount of Tris carbonate to the reference standard serum permits the carbonate dioxide content to be restored to any predetermined level upon reconstitution of the serum by the addition of distilled water, and as determined by the concentration of the Tris carbonate, depending upon the particular analytical procedure in which the reference standard serum is to be used.

The freeze-dried serum used in the practice of the invention can be prepared by any conventional method of freeze-drying whereby human serum is dried from a frozen state under high vacuum; wherein ice, or other frozen solvents, rapidly sublime to yield a porous solid. The resulting freeze-dried reference standard serum has been found to remain stable for over two years at temperatures ranging from 2 C. to C. In the practice of the invention, a quantity of Tris carbonate is added to the serum prior to freeze-drying; and the freeze-dried serum is subsequently reconstituted by the addition of distilled water prior to use. Alternately, Tris itself is added to the serum and carbon dioxide gas is bubbled into the serum prior to freeze-drying.

This reconstituting of the serum is necessary, as explained above, in order to increase the carbon dioxide content of the reconstituted serum to a predetermined level. as determined by the particular analytical procedure in which the serum is to be used for calibrating purposes. The reconstituted serum can be used directly for the calibration and control of the aforesaid apparatus.

The Tris carbonate is ideally suited as a component of the reference standard blood serum in that it does not affect in any way the precisely predetermined values of any of the constituents of the reference standard blood serum. Also, it imparts a normal pH to the serum so as to provide for the stability of its constituents, and particularly the enzymes, as mentioned above.

As explained above, the Tris carbonate serves to produce a carbon dioxide content in the reconstituted reference standard serum which is suitable for standardizing and calibrating the test equipment. As also previously stated, the Tris carbonate is colorless and has no turbid characteristics as would affect any of the tests made by the aforesaid apparatus.

The following specific example is intended to illustrate a particular reference serum illustrative of the practice of the invention.

As a first step in the preparation of the reference serum of the invention, selected samples of human blood serum were pooled. Then the Tris carbonate was added in sufficient amount to provide the desired carbon dioxide level in the reconstituted serum. For example, the Tris carbonate was formed by adding Dry Ice, as a source of carbon dioxide, to Tris. In a particular example, 65 grams of Tris was placed in 100 cc. of water, a quantity of Dry Ice was added, and the solution was allowed to stand overnight to permit the Tris carbonate to precipitate. The Tris carbonate was then dissolved in water, and added to the serum in a proportion of 7 grams of Tris carbonate per liter of serum, or in other proportions depending upon the exact level the carbon dioxide constituent of the serum is to be reconstituted. A volume of 50 milliliters of the serum and Tris carbonate mixture was then freeze-dried by a known freeze-drying process.

The freeze-dried serum, prior to use, was reconstituted by adding 50 milliliters of distilled water to a bottle containing the serum. The resulting mixture was slightly agitated by rotation of the bottle so that the serum was completely dissolved. The reconstituted pool was then analyzed by known procedures.

One specific sample of the reconstituted serum had the following constituent values which are listed herein merely by way of example:

Constituent: Value Glucose 230 mg./l00 ml. Blood urea nitrogen 69 mg./ ml. Potassium 5.4 m. eq./liter Sodium 149 m. eq./liter Carbon dioxide 30 m. eq./liter Chloride 113 m. eq./liter Alkaline phosphatase 25 KA units Transaminase (SGOT) 93 KA units Calcium 10.2 mg./l00 mil Phosphorous 4.8 mg./1()0 mil Uric acid 4.2 mg./l00 mil Cholesterol 200 mg./l00 mil Total protein 6.6 gm./l00 mil Albumin 3.9 gm./l00 mil Bilirubin 2.0 mg./l00 mil Lactic dehydrogenase 220 Wacker units What is claimed is:

1. A freeze-dried blood serum reference standard comprising blood serum having constituents of known values and a carbonate or bicarbonate Tristhydroxymethyl) aminomethane.

2. The blood serum of claim 1, in which the concentration of the aforesaid aminomethane in the serum is such that when the serum is reconstituted, the carbon dioxide content of the reconstituted serum is in a range extending from approximately 12 to approximately 35 m. eq./liter.

3. The blood serum of claim 1, reconstituted by the addition of distilled water.

4. The method of providing an accurate blood serum reference standard having a standard carbon dioxide content for use in multi-automated analytical procedures which comprises reconstituting freeze-dried blood serum containing a quantity of a carbonate or bicarbonate of Tris(hydroxymethyl)aminomethane with distilled water so that the carbon dioxide content of the reconstituted serum may be increased to a predetermined level.

5. The method set forth in claim 4, in which the concentration of the aforesaid aminomethane is such that the carbon dioxide content of the reconstituted serum is in a range extending from approximately 12 to approximately 35 m. eq./liter.

References Cited UNITED STATES PATENTS 3,466,249 9/1969 Anderson 252-408 JOHN T. GOOLKASIAN, Primary Examiner M. E. MCCAMISH, Assistant Examiner US. Cl. X.R. 23-230 B

Referenced by
Citing PatentFiling datePublication dateApplicantTitle
US3859049 *Feb 1, 1974Jan 7, 1975Marbach Edward PBlood reference standard and process for blood gas test
US3897363 *Aug 10, 1973Jul 29, 1975Baxter Laboratories IncBlood control standard
US3977995 *Oct 17, 1974Aug 31, 1976Baxter Laboratories, Inc.Calibrating fluid for blood cell counting and hemoglobin determination
US4020006 *Aug 14, 1975Apr 26, 1977Icl/ScientificFluid containing dispersed particles simulating leukocytes and method for producing same
US4054488 *Feb 26, 1976Oct 18, 1977Marbach Edward PPreservation of glucose in blood samples
US4056484 *Dec 29, 1975Nov 1, 1977Behringwerke AktiengesellschaftStable blood plasma, process for preparing it and its use as comparative plasma in coagulation tests
US4121905 *Aug 22, 1977Oct 24, 1978Jonas MaurukasProcess for preparing biological compositions for use as reference controls in diagnostic analyses
US4141856 *May 9, 1977Feb 27, 1979Dorwart Jr William VReference material for establishing anion concentrations in analytical chemistry tests
US4158544 *Jul 17, 1978Jun 19, 1979Beckman Instruments, Inc.Process for preparing a biological composition for use as a reference control in diagnostic analysis
US4299728 *Jul 21, 1980Nov 10, 1981Instrumentation Laboratory Inc.Blood gas control
US4323478 *Dec 19, 1979Apr 6, 1982Baxter Travenol Laboratories, Inc.Novel particulate compositions
US4324685 *Mar 21, 1980Apr 13, 1982Beckman Instruments, Inc.Blood serum reference standard
US4325832 *Jan 21, 1980Apr 20, 1982Beckman Instruments, Inc.Enzyme reference composition
US4369127 *Mar 9, 1981Jan 18, 1983Instrumentation Laboratory Inc.Blood gas control
US4448888 *Aug 24, 1982May 15, 1984Bio-Rad Laboratories, Inc.Compositions and method for correcting temperature dependence of hemoglobin assays
US4716119 *Oct 7, 1986Dec 29, 1987Boehringer Mannheim GmbhControl serum and process therefor
US4806486 *Jan 28, 1986Feb 21, 1989Stichting GastransportCalibration liquid for ion-specific electrodes, process for preparing them and their application
US4871678 *Apr 24, 1986Oct 3, 1989Beschrankter HaftungAgent and process for the determination of calcium
US4956299 *Dec 31, 1986Sep 11, 1990Ciba Corning Diagnostics Corp.Complement stabilization
US5238845 *Jul 29, 1992Aug 24, 1993Instrumentation Laboratory S.P.A.Method for producing a co2 calibration liquid
US5385844 *Apr 22, 1993Jan 31, 1995Boehringer Mannheim CorporationPolymer containing control reagents and polymers useful in control reagents
US6268481Jul 22, 1997Jul 31, 2001Medical Analysis Systems, Inc.Covalently coupled troponin complexes
DE2723070A1 *May 21, 1977Dec 15, 1977Dorwart JunBezugspraeparat zur festlegung einer anionenkonzentration bei chemischen analysen
DE2856988C1 *Aug 22, 1978Jan 26, 1984Jonas MaurukasVerfahren zur Herstellung von biologischen Zusammensetzungen fuer die Verwendung als Bezugs-Kontrollproben bei der diagnostischen Analyse
EP0016573A1 *Mar 4, 1980Oct 1, 1980Beckman Instruments, Inc.Enzyme reference composition
WO1979000106A1 *Aug 22, 1978Mar 8, 1979J MaurukasProcess for preparing biological compositions for use as a reference control in diagnostic analysis
U.S. Classification436/16, 436/14, 436/15, 436/11, 436/12, 436/13, 436/18, 436/19
International ClassificationG01N33/96
Cooperative ClassificationG01N2496/05, G01N33/96
European ClassificationG01N33/96
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Jul 14, 1988ASAssignment
Effective date: 19880628
Nov 2, 1982ASAssignment
Effective date: 19820917
Nov 2, 1982AS02Assignment of assignor's interest
Effective date: 19820917