Morphology antibody chromosome technique for determining phenotype and genetic status of the same cell.
The morphology antibody chromosome (MAC) technique is a combination of methods that permits analysis of both phenotypic and genetic evaluation on a single interphase or mitotic cell as a basis for lineage analysis of neoplastic and normal cells. This unit describes MAC with sequential phenotypic analysis using antibody and an alkaline phosphatase anti-alkaline phosphatase (APAAP) complex and genotypic analysis using in situ hybridization with either enzymatic or fluorescence detection. Alternate methods for phenotypic analysis are also described, which include the use of horseradish peroxidase-conjugated antibodies, fluorochrome-conjugated antibodies, May-Grunwald-Giemsa cytochemical staining, and Sudan black B cytochemical staining. An additional protocol describes G- and C-banding as alternatives to in situ hybridization (ISH) for genotyping MAC specimens. Support protocols describe methods for preparing specimens, cytospin preparations, in situ cultures, paraffin-embedded or cryostat sections, and blood and bone marrow smears. Also described is a procedure for chromosome painting of previously GTG-banded slides. An additional protocol is included for FISH analysis on sorted hematopoietic stem cells and its application in the detection of leukemic stem cells. For overcoming the drawbacks of scarcity and variability from case to case of malignant plasma cells in multiple myeloma FISH analyses, a protocol is included for the enrichment of plasma cells by immunomagnetic beads.
University of Helsinki and Helsinki University Central Hospital, Helsinki, Finland.
Sakari
S
Knuutila
Satu
S
Mustjoki
D000469:Alkaline Phosphatase / Q000276:immunology; D002871:Chromosome Banding / Q000379:methods; D020223:Chromosome Painting; D005821:Genetic Techniques; D005838:Genotype; D006801:Humans; D007158:Immunologic Techniques; D017404:In Situ Hybridization, Fluorescence / Q000379:methods; D010641:Phenotype
D000469:Alkaline Phosphatase