Immunogold labelling technique is a new type of immunolabeling technique applied to antigen-antibody reaction using colloidal gold as tracer. Colloidal gold is made of chloroauric acid under the action of reducing agents such as white phosphorus, ascorbic acid, sodium citrate and ellagic acid, etc., which polymerizes into gold particles of a specific size, and due to electrostatic effect becomes a stable colloidal state, so it is called colloidal gold.

Operation method

basic program

Principle

Utilizing the negatively charged nature of colloidal gold under alkaline conditions, it forms a strong bond with the positively charged groups of protein molecules by electrostatic attraction. In addition to antibodies, colloidal gold can also bind to a variety of other biological macromolecules, such as SPA, PHA, ConA and so on.

Materials and Instruments

Tissue Samples
Immunogold markers
Incubator Microscope

Move

1. Wet paper towels are placed on the bottom of the slide box to make a humidified box, and the slides containing the sections are taken out of the cryostat and placed into the slide box (6 slides on each side) or into the humidified box (the slides should not touch each other).2. Allow the slide to reach room temperature before it dries and spread PBS on the section (do not overflow the slide).3. Dilute the first Hangar at 4°C by microcentrifugation at 13,500 g for 2 min (40-50 μl of antibody per slide should cover the section).4. Using a Pasteur pipette attached to the pump, aspirate the PBS from the slide at one end of the section and add the antibody from the other end, cover the humidor and incubate for 1 h at room temperature.
5. PBS the slide 3 times (5 min/times), add new PBS buffer from one end of the slide and aspirate the old buffer from the other end.6. The diluted secondary antibody is centrifuged at 13,500 g for 2 min at 4°C in a microcentrifuge (40-50 μl of antibody per slide).7. Add immunogold-labeled secondary antibody and incubate for 2 hours.8. Observe results under a microscope or store in a closed slide box at room temperature.

9. Observe results under a microscope or store in a slide box at 4°C.

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