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HLA antibody specificity detecting method, cell dish and reagent kit

A detection method and specific technology are applied in the field of HLA antibody specific detection method and cell disk and kit, which can solve the problems of increasing difficulty in matching kidney sources, difficulty in finding out HLA antibody specificity, and high PRA value in the serum to be tested. To achieve the effect of good specificity and low cost

Active Publication Date: 2010-04-21
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this method is also limited by antigen overlap in natural cells. When the HLA antibodies in the serum to be tested are relatively complex and the presence of high-frequency HLA antigens, these two factors will lead to a very high PRA value in the serum to be tested. When it reaches a certain When the value is above 80%, it is difficult to find out the specificity of HLA antibody
For this part of patients with high PRA values, it takes more time and cost to use multiple detection methods to check the specificity of HLA antibodies. If kidney transplantation is required, only one-to-one cross-matching can be used Methods Finding a suitable donor kidney increases the difficulty of finding a matching kidney source

Method used

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  • HLA antibody specificity detecting method, cell dish and reagent kit
  • HLA antibody specificity detecting method, cell dish and reagent kit
  • HLA antibody specificity detecting method, cell dish and reagent kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0082] Example 1. Adjustment of HLA Antigen Frequency for Selected Donor Cell Catalogs

[0083] Table 3 HLA antigen adjustment scheme of the present invention

[0084] Active adjustment of antigen types

Frequency of HLA antigens in Chinese population (%)

Adjusted HLA antigen frequency in the cell plate (%)

w4

66

50-55

w6

62

50-55

A1

21

10-15

A2

54

25-30

A3

22

15-19

A11

33

5-15

A24

33

10-20

C7

44

25-30

w9

30

20-25

[0085] Antigen frequency adjustment principle:

[0086] In addition to the HLA antigen frequencies listed in Table 3, the selected donor cell HLA antigens also need to cover all the antigens listed in Table 1, and each antigen appears at least twice.

Embodiment 2

[0087] Embodiment 2 makes cell dish

[0088] Step 1. Select a cell donor:

[0089] Among normal healthy adults, volunteers are randomly selected to donate peripheral blood, and a total of 150 to 250 volunteers are required as the source of candidate cell donors.

[0090] Cell donor HLA tissue type identification, numbering and recording of cell HLA antigen type.

[0091] According to the HLA antigen matching of the donors, the data in Table 3 in Example 1, and the antigen frequency adjustment principle, 96 donors were selected, and 100 ml of venous peripheral blood were extracted respectively.

[0092] Step 2. Using Ficoll-Hypaque to obtain a mononuclear cell layer (PBMC), which contains a large number of peripheral blood lymphocytes. The cells were washed twice with phosphate buffered saline (PBS), and the cells were counted. (please adjust the format)

[0093] Step 3. Ninety-six donor peripheral blood lymphocytes were added to wells of a 96-well cell culture plate, with ...

Embodiment 3

[0104] Example 3. Detection of HLA antibody specificity

[0105] 1. Thaw two cell discs prepared in Example 2 at room temperature (22° C.), one cell disc is used as a base value detection disc, and the other cell disc is used as a sample detection disc. When testing multiple sera at one time, only one base test plate is needed.

[0106] 2. Wash the plate once with RPMI-1640 solution containing 10% FCS, and then wash the plate once with flow cytometer washing buffer.

[0107] 3. Add 25 μl of positive control serum to one well of the base value test plate and the sample test plate.

[0108] 4. Add 25 μl of negative control serum to the remaining 95 wells of the base value detection plate.

[0109] 5. Add 25 μl of negative control serum to another well of the specimen testing plate, and add 25 μl of patient serum to be tested into the remaining 94 wells.

[0110] 6. Incubate all base value detection plates and sample detection plates at 4°C for 30 minutes.

[0111] 7. Wash th...

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Abstract

The invention relates to an HLA antibody specificity detecting method, a cell dish and a reagent kit, belonging to the field of immunity detection. The HLA antibody specificity detecting method comprises the following steps: (1) enabling serum to be detected to respectively undergo antibody antigen reaction with peripheral blood lymphocytes taken from different donors; (2) detecting the result of the antibody antigen reaction; and (3) processing data to obtain the HLA antibody specificity. The HLA antibody specificity detecting method is characterized in that the HLA antigen frequency is adjusted by selecting the taken donors so that the detected PRA value is lower than the PRA value of the serum to be detected, and the limitation to the HLA antibody specificity detection caused by the HLA antigen overlapping phenomenon is reduced. The method can be used for detecting the HLA antibody specificity of a serum with higher PRA value and the HLA antibody specificity of the serum to be detected once so that the HLA antibody specificity detecting method is stepped forwards in a leap way and brings benefit to people.

Description

technical field [0001] The invention relates to a medical detection method and a reagent, in particular to a specific detection method for HLA antibodies, a cell plate and a kit. Background technique [0002] In clinical medicine, some diseases can cause kidney disease. When a patient's kidney disease develops to the point where he can only rely on dialysis to maintain his life, it is called end stage renal disease (ESRD). Kidney transplantation is an effective long-term treatment for patients with end-stage renal disease. In the study of organ transplantation, people found that the antigens related to transplantation are HLA-I and -II antigens, as shown in Table 1, which are the five HLA gene loci that need to be matched and detected in the current HLA matching. HLA antigens: HLA antigens at HLA-A, -B, -Cw, -DR, -DQ sites (R.Holdsworth, C.K.Hurley, S.G.E.Marsh, M.Lau, H.J.Noreen, J.H.Kempenich, M.Setterholm and M .Maiers, The HLAdictionary 2008: a summary of HLA-A, -B, -C...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/543
Inventor 才新
Owner 才新
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