Preparation of Amniocytes for Interphase Fluorescence In Situ Hybridization (FISH)

Abstract

Although FISH has been used to clarify deletions or structural rearrangements, recent work has focused increasingly on its applications to interphase analysis. This unit describes preparation of uncultured amniotic fluid cells for FISH analysis. Cells are swollen, then slides are prepared using either a cytocentrifuge or standard methods. These are then fixed and permeabilized for subsequent FISH. An Alternate Protocol describes attachment of amniocytes to a glass or plastic surface followed by hypotonic swelling, fixation, and permeabilization for subsequent FISH. Interphase FISH analysis of amniotic fluid cells is also described.

FnCas9-based CRISPR diagnostic for rapid and accurate detection of major SARS-CoV-2 variants on a paper strip

“…However, probe-based qRT PCR assays rely on amplification of a target gene with real-time analysis of DNA copy numbers and are generally not suited for genotyping point mutations associated with novel CoV2 variants ( Afzal, 2020 ; Yin, 2020 ; Vandenberg et al, 2021 ). Although strategies such as Amplification Refractory Mutation System-quantitative PCR (ARMS-qPCR) have been described in the literature for genotyping mutations in nucleic acids, these require substantial design and validation for reproducible readouts and hence are not routinely used for clinical diagnosis ( Little, 2001 ). Similarly, antigen-based SARS-CoV-2 assays which have lower sensitivity than qPCR have not yet been reported to specifically identify mutant variants.…”