A rapid, semiautomatic system of microchemical analysis for the clinical chemistry laboratory has been proposed. Five basic elements of this system are: (1) The use of siliconated-heparinized plasma. (2) The use of the calibrated-pipet-tip buret technic for measuring small (0.10-ml.) samples. (3) The use of the decantation principle as a precision step in making quantitative transfers. (4) The use of automatic syringe pipets for adding constant volumes of reagents, (5) The use of specific enzymatic methods, whenever these are applicable, for the determination of biologic constituents. The analytic system has already been applied to the determination of such important biologic constituents as glucose, urea nitrogen, phosphorus, acid and alkaline phosphatases, sodium and potassium, calcium, and total protein. The semiautomatic system permits the use of microprocedures in a clinical chemistry laboratory by persons of limited technical skill.

There are three general categories of procedures employed for the quantitative determination of the gamma globulin content of biological fluids. These are (1) electrophoretic methods, (2)

Previous reports from this laboratory 1-5 described the microdetection of sugars and other compounds containing readily reducible groups by means of tetrazolium salts. The solutions of the quaternary ammonium salts are colorless, soluble in water, and on reduction yield highly colored substances called formazans which are intensely colored and insoluble in water. SN\SN\ .RsOwing to this property and the low reduction potentials of a number of these salts, they have found use in the detection and study of reducing enzyme systems and in the detection and estimation of reducing Runctions oR organic compoundsl-L The number oR tetrazolium salts reported from this laboratory is about 35. It has been established that it is the tetrazolium ion which acts as an oxidant in alkaline media and that the threshold of stability for 2,3,5-triphenyl tetrazolium chloride is at pg 12.454. Most tetrazolium salts are stable at this pm but a number are not. Of the tetrazolium salts which have been studied, two have been selected which give readily * Dedicated to Professor Hans Lieb on occasion of his 70th birthday. biphenyl tetrazolium violet (2,5-diphenyl-3-o-biphenyl tetrazolium chloride); the other is p-anisyl tetrazolium blue (2,2'-diphenyl-5,5'-p-anisyl-3,3'-dimethoxy-4,4'-biphenylene ditetrazolium chloride). By means of the p-anisyl tetrazolium blue it is possible to detect 1/~g/ml of many reducing sugars, whereas the sensitivity of the triphenyl tetrazolium salt is 7 to I0 #g/ml. The use of the triphenyl tetrazolium salt for the determination of reducing sugars in blood was reported by Fairbridge, Willis and Booth s and by Skupp and Cheronis from this laboratory 9. The results of both groups of investigators indicated that the method based on t, he use of 2,3,5-triphenyl tetrazolium chloride was not reliable at low concentrations of glucose. The present paper reports a critical study of the quantitative determination of glucose by means Of 2,3,4-triphenyl tetrazolium chloride and the development of a method based on the use of p-anisyl tetrazolium blue, which permits the estimation of reducing sugars at very low concentrations. Determination o/ Glucose by Means o/ Tetrazolium Salts

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