Principle

Conventional staining techniques are used to uniformly stain chromosomes and leave the centromeres constricted, thus enabling the measurement of chromosome length, centromeric position, and arm ratio.

Background

Prior to 1960, when Moorehead and Nowell described the use of Giemsa in their chromosome preparations, conventional cytologic stains such as acetoorcein, acetocarmine, gentian violet, hematoxylin, Leishman's, Wright's, and Feulgen stains were used to stain chromosomes. The Romanovsky dyes (which include Giemsa, Leishman's, and Wright's stain) are now recommended for conventional staining, because the slides can be easily destained and banded by most banding procedures. Orcein-stained chromosomes cannot be destained and banded; therefore, orcein is generally not used in routine chromosome staining. Giemsa stain is now the most popular stain for chromosome analysis (Gustashaw, 1991).

Solutions

• Giemsa stain (Gurr's, Biomedical Specialties cat. # 35086)
  
• pH 6.8 phosphate buffer (Gurr's tablets, Biomedical Specialties cat. # 33199)
  
• Working stain: 4 mL Giemsa; 96 mL pH 6.8 buffer
  

Procedure

1. Place slides in a Coplin jar or staining dish.

2. Prepare the working stain and pour it over the slides.

3. Stain for 7 minutes.

4. Rinse slides in two changes of distilled water.

5. Air dry slides; mount them with a cover slip if desired. (If sequential banding procedures are to follow, coverslipping is not recommended.)

References

Gustashaw, KM. Chromosome Stains. In The ACT Cytogenetics Laboratory Manual, Second Edition, edited by M. J. Barch. The Association of Cytogenetic Technologists, Raven Press, Ltd., New York, 1991.

Primate Cytogenetics Network