Preparation of single cells from solid tissues for analysis by PCR.
This unit details a protocol for the separation of solid tissues into single cell suspensions for subsequent analysis of nucleic acids and protein. Tissues are fixed in situ by perfusion, which terminates cell processes and thus changes that would accompany dissociating the living tissue. The procedure is particularly useful when the cell type of interest represents a minor population relative to other cells types in the tissue. Once separated, individual cells or groups of a particular cell type can then be analyzed using PCR strategies. The procedure can also be adapted to allow the quantification of the number of cells within a tissue containing specific nucleic acid sequences, for example, a particular viral DNA or RNA sequence.
Sawtell
N M
NM
Children's Hospital Medical Center, Cincinnati, Ohio, USA.
D000818:Animals; D002469:Cell Separation / Q000295:instrumentation / Q000379:methods*; D004247:DNA / Q000302:isolation & purification; D007202:Indicators and Reagents; D007763:Lac Operon; D051379:Mice; D009474:Neurons; D016133:Polymerase Chain Reaction / Q000379:methods*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D016707:Tissue Fixation / Q000379:methods; D001616:beta-Galactosidase / Q000032:analysis
D007202:Indicators and Reagents; D004247:DNA; D001616:beta-Galactosidase