HU187728B - Reagent solution comprising additive, for determining erythrocites - Google Patents

Abstract

The invention relates to a reagent for erythocyte measurements, in particular for erythocyte counting. Their goal is to ensure a sufficient accuracy of Erythozytenmessungen and a high long-term stability of the measuring apparatus and the method with low equipment and process costs. The invention had the object of finding a reagent which greatly reduces the adsorbability of erythrocytes without altering their shape. As the reagent, a propylene oxide-ethylene oxide copolymer was found. As expedient has been found: the weight ratio of the propylene oxide to the ethylene oxide = 6: 4, the total molecular weight 3000g per mole and the concentration of 5-50 mg of reagent per liter of dilute blood.

Description

The present invention relates to a reagent solution containing an additive which can be used in the determination of erythrocytes, in particular in the determination of the number and volume of erythrocytes. The above measurements are usually performed in medical laboratories for diagnostic purposes.

A prerequisite for a reliable diagnosis is that the red blood cell count and volume determination must be guaranteed to be accurate to at least 3%. This accuracy is hardly achievable with cell counters or automatic analyzers that use artificial vessels for repeated use. Even continuous equipment or single-use reactor equipment often do not reliably achieve the above accuracy.

So far, the major measurement error (which may be 30%) has been attributed to microhemolysis and red blood cell aggregation. To reduce microhemolysis, reaction vessels may be made of a special material such as polyethylene. Such special materials are expensive; the reaction vessels made from them need to be used more than once for economical use. Although this achieves a significant reduction in the measurement error, it is not sufficient. In addition, the effect of reducing measurement error is reduced after repeated use. To prevent aggregation, it is essential that the measurement is performed shortly after dilution of the blood sample. However, this time may not be shorter than the reaction time required to complete the reaction between the blood elements and the reagents added to them. Therefore, the error-reducing effect of this operation is small.

According to our experiments, it was found that the measurement error is mainly caused by the adsorption processes between the red blood cells and the wall of the reaction vessel.

It is known that the main source of platelet count error is also the adsorption process. According to U.S. Patent 4,160,644, the above error can be significantly reduced by adding a solution of polyethylene glycol to the platelet suspension or the solvent. The effect can now be achieved by adding a small amount of material. Attempts to reduce the erythrocyte measurement error with the above solution have failed, even if the solution is used at higher concentrations.

Thus, we have sought a solution that can ensure, with low instrumentation and procedure costs, the accuracy of erythrocyte measurement and the stability of the measuring apparatus and measurement procedure over time.

To accomplish this object, we sought an additive which, when added to a reagent solution containing neutral salts, a buffer system and optionally a preservative, commonly used to dilute blood, strongly reduces the adsorption capacity of erythrocytes without altering its erythrocyte count and .

According to the present invention, the reagent additive is an ethylene oxide-propylene oxide block copolymer. The adsorption capacity of erythrocytes is most pronounced when the ratio of propylene oxide to ethylene oxide in the copolymer is 6: 4 and the total molecular weight is about 6%. 3000 g / mol. The concentration of the reagent additive in the reagent solution is preferably 5-50 mg / liter.

The reagent solution is, for example, 100-200 mM sodium chloride, 1-10 mM potassium chloride, as a neutral salt, for example 1-10 mM dipotassium or disodium hydrogen phosphate and 0.1-2 mM sodium phosphate buffer system. or potassium dihydrogen phosphate, for example as a preservative, for example 0.5-10 mM sodium azide. The composition and concentration of the reagent solution are not critical to the invention, one or more salts may be replaced, one or more salts may be added to the solution, or one or more salts may be omitted. In all cases, the reduction of adsorption according to the invention can be achieved by adding 5-50 mg / L reagent additive.

The following example is intended to illustrate the invention in more detail.

Example

The reactant additive used is a block copolymer of 3000 g / mol of total propylene oxide and ethylene oxide in a ratio of 6: 4. 20 mg of the above reagent additive is weighed into a reagent solution of the following composition:

154.0 mmol / l sodium chloride,

4.4 mmol / l potassium chloride,

1.0 mM sodium azide,

2.6 mmol / l disodium hydrogen phosphate,

0.4 mmol / l sodium dihydrogen phosphate.

The reagent solution supplemented with the reagent additive is diluted 1: 100,000 for the erythrocyte count. The number of red blood cells in the reaction vessel made of polypropylene is determined for the diluted blood sample. The measured number of erythrocytes is 14% higher than the number of erythrocytes determined with the same ratio of reagent solution containing no reagent additive in blood from the same patient.

Without the addition of the reagent additive, some of the red blood cells are adsorbed to the wall of the reaction vessel and thus not accessible for counting. The magnitude of the error is influenced by the material of the reaction vessel, the condition of the vessel surface, and the characteristics of the blood sample being tested. The failure rate can be as high as 30% for reusable reaction vessels for automatic equipment. Because the red blood cell concentration is also used to calculate additional blood parameters, the measurement error also affects the calculated parameters. The reagent additive inhibits the adsorption of red blood cells to the surface of the reaction vessel, thereby reducing measurement error.

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Claims (2)

An additive reagent solution for erythrocyte count, containing one or more neutral salts, typically 100-200 mM sodium chloride, 1-10 mM potassium chloride, pH 1-8, at a conventional concentration. mmol / l disodium hydrogen phosphate, 0.1-2 mmol / l sodium dihydrogen phosphate and optionally a preservative, preferably 0.5-10 mmol / l sodium azide. that the additive is propylene oxide ethylene oxide block copolymer at a concentration of 5-50 mg / L. 2. The reagent side of claim 1, wherein the block copolymer has a propylene oxide to ethylene oxide weight ratio of from 5 to 7: 35 and a total molecular weight of 2000 to 4000 g / mol. Without illustration