Biomedical Engineering Reference
In-Depth Information
5
In-Cell Polymerase Chain Reaction
Strategy and Diagnostic Applications
T. Vauvert Hviid
Summary
In situ
polymerase chain reaction (PCR) refers to the amplification of specific nucleic
acid sequences and subsequent visualization of the PCR products in tissue sections. When
PCR is performed in fixed cells in suspension the term
in-cell PCR
is normally applied
instead. For example, in-cell PCR has been used for intracellular detection of specific
viruses. Furthermore, in mixtures of cells in suspension with different genotypes, in-cell
PCR may be used for intracellular detection of DNA or mRNA sequences specific for
one of the genotypes/population of cells. An in-cell PCR method is described that makes
it possible to genotype a specific gene of interest derived from one individual in a mix-
ture of cells from two individuals. This new method is based on in-cell PCR and depen-
dent upon the presence of a well characterized and specific DNA sequence or
polymorphism, which has to be present only in the cells from the individual being
genotyped for the gene sequence of interest. The in-cell PCR method might find diag-
nostic applications in detection of intracellular pathogenic sequences in subgroups of
cells and in noninvasive prenatal genetic diagnoses with the use of the few fetal cells
circulating in maternal blood.
Key Words:
In-cell PCR;
in situ
PCR; genotyping; cell mixtures; fetal cells; mater-
nal blood; prenatal diagnosis.
1. Introduction
In situ
polymerase chain reaction (PCR) refers to the use of the PCR method
for amplification of specific nucleic acid sequences and subsequent visualiza-
tion of the PCR products in tissue sections. In this way, the recognition of the
presence of specific mRNA or DNA sequences in different types of cells and
tissues can be obtained through the combined examination of PCR/reverse-
transcription (RT)-PCR product signal and histology. When PCR is performed
in fixed cells in suspension the term
in-cell PCR
is normally applied instead.
