Immunocytochemistry

We use Immunocytochemistry to investigate protein-complexes.

Acrylic resins:

1) LR White.

2) Lowicryl K11M.

 

 

Workflow – 1:

1) Chemically fix tissue cells with paraformaldehyde solution.

2) Infiltrate in a variety of resin/acetone mixtures.

3) Polymerise in LR White at -20°C or Lowicryl K11M at -60°C under UV irradiation.

4) Cut the ultra section at room temperature.

5) Block with BSA buffer solution.

6) Immuno label with 1° antibody & 2° antibody-conjugated colloidal golds on the grid.

7) Chemically fix with glutaraldehyde and wash with water.

8) TEM.

 

 

Workflow – 2:

1) Vitrificate tissue cells by HPF.

2) Place the cells in cold uranyl acetate in methanol at -90°C for several hours – days.

3) Follow same procedure as for Freeze substitution.

4) Embed cells in Lowicryl K11M and polymerise at -60°C by UV irradiation.

5) Cut the ultra section at room temperature.

5) Block with BSA buffer solution.

6) Immuno label with 1° antibody & 2° antibody-conjugated colloidal golds on the grid.

7) Chemically fix with glutaraldehyde and wash with water.

8) TEM.

 

 

Post-embedding immuno-gold labelling

Lung carcinomas

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