SU1677634A1 - Method for estimation of lymphocyte agglutination - Google Patents

Abstract

The invention relates to an immunological analysis of cells. The purpose of the invention is to increase the sensitivity of the method and reduce the analysis time. Before applying the suspension of lymphocytes before applying concanavalin A, NADH is added at a concentration of 70-300 µg / ml or catalysis at a concentration of 20-75 µg / ml, or leroxidase at a concentration of 20-75 µg / ml. Lymphocyte agglutination is determined by the intensity of the suspension's light permeability. Compared to the prototype, the sensitivity is increased 5 times, and the analysis time is reduced by a factor of b.

Description

The invention relates to the field of immunochemical analysis of cells and can be used to control biotechnological processes and in scientific research.

The aim of the invention is to increase the sensitivity of the method and reduce the analysis time.

The method is carried out as follows.

Before applying concanavalin A (kon A), NADH at a concentration of 70-300 µg / ml or catalase at a concentration of 20- 75 µg / ml, or peroxidase at a concentration of 20-100 µg / ml are added to the test cell suspension and the agglutination rate is determined by change the intensity of light transmission.

PRI me R 1. Lymphocyte suspension is prepared in Dulbecco's phosphate-buffered saline, pH 7.3 with a working cell concentration of 5-106 cells / ml and the number of viable cells determined by trypan blue, not less than 95%. At 37 ° C, NADH in concentrations of 0, 65, 70, 100, 150, 200, 250, 300, 305 µg / ml, respectively, and concanavalin A (con A) from Sigma (USA) at a concentration of 25 µg / ml. The intensity of light transmission is measured on a laser nephelometer.

Addition of NADH to the test sample increases the rate of agglutination with a minimum content of the agglutination agent con A. The light transmission can be recorded.-Ic can be performed after 5 min after the addition of NADH and con A.

Example 2 Peroxidase at concentrations of 0, 15, 20, Z, 50, 70, 100 and 105 µg / ml, respectively, and end A at a concentration of 25 µg / ml, are added to the lymphocyte suspension prepared in Example 1, then measured light transmission intensity

Example 3 To a suspension of lymphocytes prepared according to Example 1, catalase is added in concentrations of 0, 15. 20, 30, 50

WITH

to

Xi

Xj

ABOUT

Ca

-1's

.,BUT

50, 60.75, 80 µg / ml, respectively, and con A at a concentration of 25 µg / ml, then the light transmission intensity is measured.

Thus, the proposed method has the following advantages compared with the prototype.

By reducing the agglutination time: with a minimum concentration of the agglutination agent con A, the addition of NADH, catalase or peroxidase increases the agglutination rate by more than 6 times (the agglutination in the proposed method is recorded 5 min after the addition of con A, and the prototype after 30 min).

Greater sensitivity: an equal time to reach lymphocyte agglutination is achieved in the prototype with

25 µg / ml con a, and in the proposed method at 5 µg / ml.

Claims (1)

Invention Formula Method of determining agglutination lymphocytes, comprising adding concanavalin A to the lymphocyte suspension, followed by incubation and agglutination evaluation, characterized in that In order to increase the sensitivity of the method and reduce the time of analysis, addition of concanavalin A is added to a suspension of lymphocytes additionally Nicotin dine nucleotide at a concentration of 70300 µg / ml or catalase at a concentration of 20-75 µg / ml or peroxidase at a concentration of 20-100 µg / ml and agglutination is determined by the intensity of the light transmission of the suspension.