Tissue microarray immunofluorescence (IF) single labeling

Service introduction:

Immunofluorescence technique is a technique established on the basis of immunology, biochemistry and microscopy. The method of using fluorescent antibodies to trace or examine corresponding antigens is called the fluorescent antibody method; the method of using known fluorescent antigen markers to trace or examine corresponding antibodies is called the fluorescent antigen method. These two methods are collectively referred to as immunofluorescence technique. However, in practice, the fluorescent antigen technique is rarely used, so the fluorescent antibody technique is often referred to as immunofluorescence technique.

Labeling a single antigen on one section is referred to as immunofluorescence (IF) single labeling, which usually can label red or green fluorescence, and nuclei are labeled with DAPI. Tissue microarray immunofluorescence allows dozens to hundreds of small tissues to be neatly arranged on a single slide for staining, so that the experimental conditions of tissue samples on the microarray remain consistent, saving time and reagents, and facilitating unified analysis after scanning. Compared with single-section staining, both time and cost can be reduced; the disadvantage is that the selected tissue area is relatively small and cannot comprehensively reflect the expression status of individual samples. The tissue microarray immunofluorescence workflow is basically the same as that of conventional tissue immunofluorescence, but because the samples are relatively precious, the operating procedures are more stringent.

Main applications:

Immunofluorescence staining and imaging analysis are indispensable detection techniques for studying tissue morphology and in situ antigen expression in tissues, and are widely used in clinical pathology and various fields of medical and biological research. Any substance that can act as an antigen or hapten, such as proteins, polypeptides, amino acids, polysaccharides, phospholipids, enzymes, hormones, pathogens and receptors, can be detected at the cellular and subcellular levels.

Detection principle:

Based on the principle of specific antigen-antibody binding, immunofluorescence technique first labels known antibodies (or antigens) with fluorophores, and then uses these fluorophore-labeled antibodies (or antigens) as probes to detect the corresponding antigens (or antibodies) in fixed cells or tissues. When observed under a fluorescence microscope, the fluorophore in the antigen-antibody complex emits fluorescence of a certain wavelength after being irradiated by excitation light, thereby determining the nature and localization of the target antigen (or antibody). Quantitative techniques (such as flow cytometry) can also be used to determine the content.

Experimental workflow:

Deparaffinization and rehydration - antigen retrieval - serum blocking - primary antibody incubation - secondary antibody incubation - DAPI nuclear staining - mounting - microscopic examination.

Sample requirements and shipping conditions:

1. Provide properly fixed and qualified samples. After tissue excision, immediately fix the tissue with an appropriate fixative. The volume of fixative is recommended to be greater than 10-15 times the tissue volume. Fresh specimens should be fixed in appropriate containers for 24-48 h; large specimens should be fixed for 12 h, then cut and further fixed. Samples should be fixed at room temperature (around 24°C) or refrigerated (4°C); do not freeze. Fixed samples should be transported at room temperature for submission.

2. Smears and coverslip preparations must be fully fixed, and paraffin sections must be thoroughly deparaffinized before the experiment.

3. Provide accurate antibody information.

Precautions: