Hematoxylin: A Simple, Multiple-Use Dye For Chromosome Analysis
Hematoxylin: A Simple, Multiple-Use Dye For Chromosome Analysis
Hematoxylin: A Simple, Multiple-Use Dye For Chromosome Analysis
chromosome analysis
Marcelo Guerra
Departamento de Botânica, Centro de Ciências Biológicas, Universidade Federal de Pernambuco, Cidade
Universitária, 50670-420 Recife, PE, Brasil.
ABSTRACT
A staining mixture of hematoxylin-iron alum combined with a strong hydrochloric
hydrolysis was successfully applied for chromosome observation of several kinds of
plants and some animals. Slightly different procedures were developed for different
materials and objectives. For plant cells, the most important technical aspect was
the use of 5 N HCl hydrolysis, which resulted in a very transparent cytoplasm,
combined with an intense, specific hematoxylin stain. This technique is
recommended for cytogenetical analysis in general, and it is especially indicated for
practical classes, due to its simplicity and high reproducibility of results. Moreover,
the deep contrast observed makes this technique very useful for sequential staining
of cells previously analyzed with other stains, as well as for materials with fixation
problems.
INTRODUCTION
Hematoxylin, combined in the proportion of 4:1 with alum,
was one of the first staining solutions used to analyze
chromosomes, and for a long time it was among the stains
preferred by classic cytogeneticists. Hematoxylin, itself, is
not a dye. First, it needs to be oxidized, either by alum or
by employing a tissue mordant. L. Taits, as early as 1875,
drew attention to hematoxylin's capacity for preferentially
staining the nucleus (Conn, 1929).
Methods
1 - Staining-followed-by-squashing method
2 - Staining-after-squashing method
A - Plant chromosomes
B - Animal chromosomes
3 - Sequential staining
ACKNOWLEDGMENTS
The author is very grateful to Dr. Maria José Lopes for supplying cytological
material of bats and grasshoppers, Ana Emilia Barros and Silva for technical
assistance and to his students Leonardo Félix, Gianna Carvalheira and André
Vanzela for several tests and comments. This work was supported by CNPq, BNB
and FACEPE.
RESUMO
Uma mistura corante à base de hematoxilina e alúmem férrico, combinada com
uma hidrólise clorídrica forte, foi utilizada na observação de cromossomos de
diversos tipos de vegetais e alguns animais, sempre com bons resultados.
Procedimentos ligeiramente diferentes foram desenvolvidos para diferentes
finalidades e diferentes materiais. No caso das células vegetais, o aspecto técnico
mais importante foi o uso da hidrólise clorídrica, que torna o citoplasma muito
transparente, combinado com a coloração intensa e muito específica produzida pela
hematoxilina. A técnica é recomendada para análise citogenética em geral, sendo
especialmente indicada para aulas práticas, devido à simplicidade dos
procedimentos e à alta repetibilidade dos resultados. Além disso, devido ao maior
contraste obtido entre citoplasma e cromatina, ela se mostrou muito útil para a
coloração seqüencial de células previamente analisadas com outros corantes e para
materiais com problemas de fixação.
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