Intended Use
For In Vitro Diagnostic Use
Summary and Explanation
MART-1/Melan-A is a putative 18 kDa transmembrane protein consisting of 118 amino acids. It has a single transmembrane domain. MART-1/ Melan-A is a protein antigen found on melanocytes. Antibodies against this antigen are used to recognize cells of melanocytic differentiation, useful for the diagnosis of Melanoma. The same name is used to refer to the gene which codes for this antigen.
The MART-1/Melan-A antigen is specific for the melanocyte lineage found in normal skin, retina, and melanocytes, but not in other normal tissues. It is thus useful as a marker for Melanocytic Tumors, with the caveat that it is normally found in benign nevi as well. This antibody is very useful in establishing the diagnosis of Metastatic Melanomas.
Presentation
Anti - MART-1 is a mouse monoclonal antibody derived from cell culture supernatant that is concentrated, dialyzed, filter sterilized and diluted in buffer pH 7.5, containing BSA and sodium azide as a preservative.
Mohs IHC Procedure
Specimen Preparation of Mohs Frozen Tissues
- Embed the specimen in OCT inside a cryostat.
- Cut sections at 4-5 µm and mount on a positively charged glass slide such as the Bio SB Hydrophilic Plus Slides (BSB 7028) or in the lower third of the TintoDetector Cap Gap slides (BSB 7006).
- Air dry the slide at room temperature for 2 minutes and then incubate the slide at 60 °C for 3 minutes in an incubator or dry bath.
- Fix in 100% acetone for 2 minutes at room temperature, and let the slide air dry.
IHC Detection Procedure
- Transfer slides to ImmunoDNA washer TBST or PBST buffer.
- For manual staining, perform antibody incubation at ambient temperature. For automated staining methods, perform antibody incubation according to instrument manufacturer’s instructions.
- Wash slides with ImmunoDNA washer, TBST, PBST Buffer or DI water.
- Continue IHC detection protocol. Wash slides between each step with ImmunoDNA washer or Tris or PBS Buffer solution.
Abbreviated Mohs PolyDetector DAB HRP Brown or HRP Green Immunohistochemical Protocol
- Incubate slides with Peroxidase Blocker for 30 seconds
- Wash with Buffer (TBST or PBST)
- Incubate with Primary Antibody for 4 min
- Wash with Buffer (TBST or PBST)
- Incubate with HRP Label for 3 min
- Wash with Buffer (TBST or PBST)
- Prepare
- DAB Brown (1 drop of DAB Chromogen in 1ml of DAB Buffer; mix well)
- or HRP Green (1 drop of HRP Green Chromogen in 1 ml of HRP Green Buffer; mix well)
- Incubate with DAB or HRP Green for 2 min
- Wash with Buffer (TBST or PBST)
- Counterstain with Hematoxylin or Nuclear Fast Red for 30 seconds
- Wash with Buffer (TBST or PBST)
- Mount with AquaMounter or dehydrate the tissue with Fast ChromoProtector then mount with PermaMounter
Step
|
Mohs PolyDetector HRP Green or DAB Protocol |
| Peroxidase Blocker |
0.5 min. |
| Primary Antibody |
4 min. |
| 1st Step Detection |
3 min. |
| Substrate-Chromogen |
2 min. |
| Counterstain / Coverslip |
Varies |
This protocol can also be used with FFPE tissues retrieved with Citrate or EDTA.
