I would appreciate an efficient protocol for measuring sister chromatid exchange or suggestions for a commercial kit available to purchase.
You can have a look at the online published paper of ours detailed: Detection of Genotoxicity and Mutagenicity of
Chlorthiophos Using Micronucleus,
Chromosome Aberration, Sister Chromatid
Exchange, and Ames Tests
Dilek Akyıl, Muhsin Konuk
Environmental Toxicology DOI 10.1002/tox
hoping that helps you,
regards
BrdU is a good nucleoside to study sister chromatid exchanges in proliferation cells in vivo & in vitro also. And we can easily measure the exchanges after staining. It is cheap as well as without kit protocol.
SCE Stain
Materials
1. Bromodeoxyuridine (BrdU) stock solution, 1 mg/ml in PBS, HBSS, or SF-DMEM.
2. Hoechst 33528 (bisbenzimide) stock solution, 10 mg/ml
3. McIlvaines buffer pH8
Sol.A-0.1 M Citric Acid 2.25 ml
Sol.B-0.2 M Disodium phosphate 97.75 ml
4 6% Geimsa in Gurr's Buffer (Gibco/BRL) pH6.8
Procedure
1. Add BrdU to growing cultures to a final concentration of 10 ug/ml. Incubate for 2 cell cycles, 36 to 60 hours, depending on the cell line.
2. Harvest cells for metaphase chromosomes as usual. Do not dry the slides on a slide warmer or heat.
3. Stain slides in Hoechst 33528 (200 ug/ml) in water for 30 min. At room temp. Rinse well with water and air dry or blot dry, do not heat.
4. Place 100 ul of McIlvaines buffer on the slide and cover with a 22x60 mm coverslip.
5. Place slide on slide warmer at 55-60*C under long and short wave UV light for 20 min (we use a UV crosslinker). Rinse slide with water and blot or air dry .
6. Stain in Geimsa for about 10 min.
From: Goto, K., S. Maeda, Y. Kano, T. Sugiyama. Factors involved in differential geimsa-staining of sister chromatids. Chromasoma. 66:351-359, 1978.
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