(fluorescence in situ hybridization)
Designed by OCTAVIAN HENEGARIU, Yale University
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PCR|dUTP label|FISH| FISH guide| CCK |Slide prep| CM-FISH | TM-FISH |mArrays |Home
- FISH guide and troubleshooting:is a summary of the author's practical experience with numerous FISH procedures used or tested in time. This section contains over 150 images and close to 130 manuscript pages, which detail numerous factors that influence hybridization results (signal, background). There are tips on slide preparation, probe preparation, hybridization buffers, probe mapping, multicolor FISH, CGH, M-FISH, fiber FISH, RNA in situ, immunostaining and many other. The most important observations detailed here were published within several of the author's previous manuscripts.
- Color changing karyotyping (CCK)(Ref: Nature Genet 23 (3):263-4 (1999) Get article here ) details a procedure allowing color karyotyping of human chromosomes without investments in new hardware or software. Any fluorescence microscope equipped with 3 filters is sufficient. Any camera currently available on your system can be used for image aquisition (digital photographic camera, CCD camera, cooled CCD camera). Image processing is done using a widely available image processing software (Photoshop, Adobe Corporation) on any computer. (Topics: 1. CCK principle; -2. CCK examples;-3. CCK methods/protocols)
- Centromeric multiplex FISH (CM-FISH)(Ref: Lab Invest. 2001 Apr;81(4):475-81. Get article here) : describes a 2-hour FISH procedure, which uses combinatorial labeling of 6 fluorophores (similar to M-FISH), to identify aneuploidies and marker chromosomes on one slide, either on metaphases or on interphase nuclei.
- Telomeric multiplex FISH (TM-FISH)(Ref: Lab Invest. 2001 Apr;81(4):483-91. Get article here): describes three FISH procedures, using combinatorial labeling of 3-5 different fluorophores (similar to M-FISH), to simultaneously detect all 41 human subtelomeric probes on the same slide. A similar approach for the mouse chromosomes (mTM-FISH) is also detailed.
- Identification of all mouse chromosomes using BACs(Ref: Mamm Genome 2001, Vol 12, p462-465. Get article here). A triple color FISH procedure used to detect (on the same metaphase) all mouse chromosomes, using 40 mouse-specific BAC clones. Useful for probe/gene mapping. As there are only three fluorescence colors, the procedure can can be performed using any fluorescence microscope equipped with three filters.
- DNA size measurements(Ref: Biotechniques 2001, Vol 31, p246-250. Get article here). A simple and rapid procedure for stretching DNA molecules on slides (using DNA cloned in vectors, such as plasmids, cosmids, BACs, PACs, YACs or from genomic DNA), and assessing the size of the various DNA fragments after YOYO staining.
- µArrays(DNA Microarrays analysis); guide and troubleshooting; complete protocols for hybridization, analysis.
Coming up !!
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"Happy New Millenium",with"The Dancing Chromosomes"
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