DNA Comet Giemsa Staining for Conventional Bright-Field Microscopy
نویسندگان
چکیده
This study was undertaken to evaluate the compatibility of Giemsa staining protocol with the comet assay. We showed, for the first time, that DNA comets can be visualized and analyzed using Giemsa staining. We generated DNA damage dose response curves for human peripheral blood lymphocytes exposed to X-ray radiation using the comet assay with either SybrGreen I or Giemsa stain. The dose response curves were fitted by linear regressions (R2>0.977). The SybrGreen I results showed only ~1.2-fold higher slope coefficient (method sensitivity) compared to the Giemsa results. The unexpectedly high sensitivity of Giemsa staining for the comet assay is due to the Romanowsky-Giemsa effect, the stain photo-stability and the higher resolution of bright-field imaging compared to fluorescence imaging. Our results demonstrate that Giemsa staining can effectively be used for measuring DNA damage by the comet assay. The low cost and availability of Giemsa stain makes this method affordable for any low budget research and will facilitate new applications of the comet assay in biology and medicine.
منابع مشابه
Early detection of Chlamydia trachomatis using fluorescent, DNA binding dyes.
HeLa 229 cells were infected with genital tract strains of Chlamydia trachomatis. After incubation for varying times the infected cells were fixed and stained with the fluorescent DNA binding dyes Hoechst 33258 or DAPI for comparison with conventional Giemsa stain. Fluorochrome-treated preparations were examined by incident ultraviolet fluorescence microscopy and the Giemsa-stained preparations...
متن کاملEarly detection of Chlamydia trachomatis using fluorescent , DNA binding dyes
HeLa 229 cells were infected with genital tract strains of Chlamydia trachomatis. After incubation for varying times the infected cells were fixed and stained with the fluorescent DNA binding dyes Hoechst 33258 or DAPI for comparison with conventional Giemsa stain. Fluorochrome-treated preparations were examined by incident ultraviolet fluorescence microscopy and the Giemsa-stained preparations...
متن کاملPlasmodium DNA contamination between blood smears during Giemsa staining and microscopic examination.
Giemsa-stained blood smears are mainly used for microscopic examination to diagnose malaria. However, they may also be subjected to polymerase chain reaction (PCR) to confirm diagnosis or for retrospective studies requiring the analysis of old smears. We investigated the possibility of DNA contamination occurring during automated Giemsa staining or due to the failure to clean the oil-immersion ...
متن کاملIntroducing a combined Leishman-Giemsa stain as a new staining technique for avian blood smears
Background: The use of conventional stains including Giemsa, Wright and Leishman have become an essential tool for differential diagnosis of cells in peripheral blood. Aims: The aim of the study was to develop a new combination of Leishman-Giemsa (L&G;) stain for avian blood smears and to compare its efficacy with conventional staining methods. Methods:<...
متن کاملBright-field microscopy visualization of proteins and protein complexes by in situ proximity ligation with peroxidase detection.
BACKGROUND The in situ proximity ligation assay (PLA) allows a protein or protein complex to be represented as an amplifiable DNA molecule. Recognition is mediated by proximity probes consisting of antibodies coupled with oligonucleotides. Upon dual binding of the proximity probes, the oligonucleotides direct the formation of a circular DNA molecule, which is then amplified by rolling-circle re...
متن کامل