Conventional HIV antibody detection method
Time：2019/06/26
The screening method
1. Enzyme-linked immunosorbent assay (ELISA)
This type of test can use blood (including serum, plasma and filter paper dried blood spots), urine samples, and ELISA is mostly HIV antibody detection reagent. The HIV antigen-antibody combined detection reagent can simultaneously detect HIV-1 P24 antigen and HIV-1/2 antibody in blood. The HIV antigen/antibody is coated on a solid phase carrier, and the sample to be tested and the enzyme-labeled HIV antigen/antibody are added, and the substrate is colored, and the result is measured by a microplate reader. Negative and positive controls for valid testing must be in accordance with the kit.
2. Chemiluminescence or immunofluorescence test (CIA/IFA)
Such assays use luminescent or fluorescent substrates, which can be used with blood (including serum and plasma), urine samples, and can be used to detect antibodies or antigen antibodies. The HIV antigen/antibody is coated on a solid phase carrier, and the test sample and the enzyme or fluorescently labeled HIV antigen/antibody are added, and a luminescent or fluorescent substrate is added, and the result is measured by a luminescence or a fluorometer. Negative and positive controls for valid testing must be in accordance with the kit.
3. Rapid test (RT) and other tests
This type of test can use blood, oral mucosal exudate, easy and fast operation, suitable for emergency testing, outpatient emergency testing, VCT and testing points. Results can generally be obtained in 10 to 30 minutes.
Gelatin particle agglutination test (PA): is a simple method for HIV antibody detection. Gelatin sensitized gelatin particles are applied to the sample to be tested. When the sample to be tested contains HIV antibodies, the gelatin particles undergo agglutination reaction with the antibody, and the results are judged according to the agglutination. There are two types of PA reagents: simultaneous detection of HIV-1 and HIV-2 antibodies and detection of HIV-1 and HIV-2 antibodies, respectively. Negative and positive control controls for effective testing are subject to the kit requirements.
Immunodiafiltration test: spot ELISA and spot immunocolloidal gold (or colloidal selenium) rapid test: both nitrocellulose membranes are used as carriers, HIV antigens are fixed in spots or lines on the membrane, and samples to be tested are used, and microporous membranes are used. The filterability allows antigen-antibody reactions. Positive results show colored spots or bands on the antigenic site of the membrane. The reaction time is within 10 minutes. The quality control points of an effective test must be developed.
Immunochromatographic assay: The nitrocellulose membrane was used as a carrier, and the HIV antigen was linearly fixed on the membrane. The sample to be tested migrated along the solid phase carrier, and the positive result showed a colored band on the antigenic site of the membrane. The quality control tape for an effective test must be colored. The reaction time is within 30 minutes.
Antibody confirmation method
1. Western blot test (WB)
WB can use blood serum, plasma and filter paper to dry blood spots. WB uses an indirect method to detect anti-HIV-1/HIV-2 specific antibodies in samples. Polyacrylamide gel electrophoresis separates HIV-1 proteins of varying molecular weights and then transfers the separated different protein bands to a nitrocellulose membrane (or PVDF membrane). The membrane was cut into strips, each of which contained an HIV antigen separated by electrophoresis. After the sample to be tested is properly diluted, it is added to the nitrocellulose membrane and shaken at a constant temperature to make it fully contact with the reaction. If the serum contains HIV antibody, it will bind to the antigen band on the membrane strip. After the addition of the anti-human-IgG enzyme conjugate and the substrate, according to the occurrence of the band, the sample to be tested is judged to be positive, negative or uncertain according to the criteria of the kit specification.
2. Band/Linear Immunization Test (RIBA/LIA)
RIBA/LIA uses an indirect method to detect anti-HIV-1/HIV-2 specific antibodies in samples. The strip of the kit is coated with different recombinant antigenic fragments of HIV-1/HIV-2. After the sample to be tested is added, the corresponding antibody reacts with the antigen specifically; then, anti-human IgG (alkaline phosphate) is added. Enzyme labeling) is combined with HIV-specific IgG antibody; after the chromogenic substrate is added, under the catalysis of alkaline phosphatase, the binding site of the specific antibody appears visible to the naked eye, and is judged according to the criteria of the kit specification. The sample is tested as positive, negative or uncertain.
3. Other methods:
Three enzyme-linked immunoassays, three rapid assays or enzyme-linked rapid assays, immunochromatography and immunodiafiltration assays can be used to confirm specific conditions.
1. Enzyme-linked immunosorbent assay (ELISA)
This type of test can use blood (including serum, plasma and filter paper dried blood spots), urine samples, and ELISA is mostly HIV antibody detection reagent. The HIV antigen-antibody combined detection reagent can simultaneously detect HIV-1 P24 antigen and HIV-1/2 antibody in blood. The HIV antigen/antibody is coated on a solid phase carrier, and the sample to be tested and the enzyme-labeled HIV antigen/antibody are added, and the substrate is colored, and the result is measured by a microplate reader. Negative and positive controls for valid testing must be in accordance with the kit.
2. Chemiluminescence or immunofluorescence test (CIA/IFA)
Such assays use luminescent or fluorescent substrates, which can be used with blood (including serum and plasma), urine samples, and can be used to detect antibodies or antigen antibodies. The HIV antigen/antibody is coated on a solid phase carrier, and the test sample and the enzyme or fluorescently labeled HIV antigen/antibody are added, and a luminescent or fluorescent substrate is added, and the result is measured by a luminescence or a fluorometer. Negative and positive controls for valid testing must be in accordance with the kit.
3. Rapid test (RT) and other tests
This type of test can use blood, oral mucosal exudate, easy and fast operation, suitable for emergency testing, outpatient emergency testing, VCT and testing points. Results can generally be obtained in 10 to 30 minutes.
Gelatin particle agglutination test (PA): is a simple method for HIV antibody detection. Gelatin sensitized gelatin particles are applied to the sample to be tested. When the sample to be tested contains HIV antibodies, the gelatin particles undergo agglutination reaction with the antibody, and the results are judged according to the agglutination. There are two types of PA reagents: simultaneous detection of HIV-1 and HIV-2 antibodies and detection of HIV-1 and HIV-2 antibodies, respectively. Negative and positive control controls for effective testing are subject to the kit requirements.
Immunodiafiltration test: spot ELISA and spot immunocolloidal gold (or colloidal selenium) rapid test: both nitrocellulose membranes are used as carriers, HIV antigens are fixed in spots or lines on the membrane, and samples to be tested are used, and microporous membranes are used. The filterability allows antigen-antibody reactions. Positive results show colored spots or bands on the antigenic site of the membrane. The reaction time is within 10 minutes. The quality control points of an effective test must be developed.
Immunochromatographic assay: The nitrocellulose membrane was used as a carrier, and the HIV antigen was linearly fixed on the membrane. The sample to be tested migrated along the solid phase carrier, and the positive result showed a colored band on the antigenic site of the membrane. The quality control tape for an effective test must be colored. The reaction time is within 30 minutes.
Antibody confirmation method
1. Western blot test (WB)
WB can use blood serum, plasma and filter paper to dry blood spots. WB uses an indirect method to detect anti-HIV-1/HIV-2 specific antibodies in samples. Polyacrylamide gel electrophoresis separates HIV-1 proteins of varying molecular weights and then transfers the separated different protein bands to a nitrocellulose membrane (or PVDF membrane). The membrane was cut into strips, each of which contained an HIV antigen separated by electrophoresis. After the sample to be tested is properly diluted, it is added to the nitrocellulose membrane and shaken at a constant temperature to make it fully contact with the reaction. If the serum contains HIV antibody, it will bind to the antigen band on the membrane strip. After the addition of the anti-human-IgG enzyme conjugate and the substrate, according to the occurrence of the band, the sample to be tested is judged to be positive, negative or uncertain according to the criteria of the kit specification.
2. Band/Linear Immunization Test (RIBA/LIA)
RIBA/LIA uses an indirect method to detect anti-HIV-1/HIV-2 specific antibodies in samples. The strip of the kit is coated with different recombinant antigenic fragments of HIV-1/HIV-2. After the sample to be tested is added, the corresponding antibody reacts with the antigen specifically; then, anti-human IgG (alkaline phosphate) is added. Enzyme labeling) is combined with HIV-specific IgG antibody; after the chromogenic substrate is added, under the catalysis of alkaline phosphatase, the binding site of the specific antibody appears visible to the naked eye, and is judged according to the criteria of the kit specification. The sample is tested as positive, negative or uncertain.
3. Other methods:
Three enzyme-linked immunoassays, three rapid assays or enzyme-linked rapid assays, immunochromatography and immunodiafiltration assays can be used to confirm specific conditions.