Do you heat the malachite green?
5 minutes with initial heating to boiling point (bunsenburner under
slide) should do the trick.
Or maybe your culture is not old enough and no spores have been formed
yet.
Loes
Diego Bonatto - 219 wrote:
> Hello people,
>> I'm working with a Bacillus sp. isolated from soil. I'm trying to
> identified the endospore of it using the Schaeffer-Fulton technique
> for light microscopic staining of endospore (green malachite for 10
> minutes and fucsine for 30 seconds). In resume I can not seen any
> endospore in microscopy using this protocol. Is this the best way
> to identify a endospore? I now that are methods more drastics, like
> to submit a culture of Bacillus at 80 degrees for 10 minutes and
> inoculate a culture media with these "cook" endospores. But I want
> a light microscopic staining method (I want to see an endospore
> inside the cell!!). If you know a method for staining endospore
> please send me a reply.
> Another question: are there good references for taxonomy of
> Bacillus? The Bergey's Manual of Determinative Bacteriology is
> very limited and I don't have the The Bergey's books of Systematic
> Bacteriology! Any reference is welcome, specially references that
> deal with fenotypic characters of Bacillus megaterium and Bacillus
> subtilis and methods for identification of these species of Bacillus.
> Thanks
> Diego Bonatto
>diego at dna.cbiot.ufrgs.br>>> Diego Bonatto
> Universidade Federal do Rio Grande do Sul
> Centro de Biotecnologia do Estado do Rio Grande do Sul
> Departamento de Biologia Celular e Molecular
> Brasil
