Antigen retrieval protocol

(necessary for paraffin section)

Protease-induced antigen retrieval (PIER)

• Antigen retrieval buffer for protease-induced antigen retrieval (epitope retrieval):

  • 1. 2 mg/ml pepsin in Tris-HCl pH 2.0
  • 2. Or 0.5 mg/ml trypsin in Tris-HCl pH 8.0

• Protease-induced antigen retrieval protocol:

  • 1. Pre-warm the buffer to 37°C;
  • 2. Keep the sections in retrieval buffer at 37°C for 10-20 minutes.

Suitable for antigens that are sheltered by fixative such as:

Collagen, GFAP, Complement, Cytokeratin, C-erB-2, LCA, LN, etc.

Heat-induced antigen retrieval (HIER)

• Antigen retrieval buffer for heat-induced antigen retrieval (epitope retrieval):

• Heat-induced antigen retrieval protocol:

  • 1. Microwave the antigen retrieval buffer until boiling and power it off, put in the sections;
  • 2. Repeat step 1 for one or two more times for every 5-10 minutes;
  • 3. Cool down at room temperature and wash with dH2O.

Preferred antigens:

Bcl-2, Bax, c-myc, E-cadherin, ChromograninA, Heatshock protein, HPV, P-glycoprotein, PKC, PCNA, etc.