7 replies to this topic

[Liang]

I’m looking for a method to separate yeast cells from small human lung tissues. Currently there are no antibodies to this yeast available. I wonder if there are chemical/physical methods to separate yeast cells from human lung tissues infected with this yeast. Thank you in advance.

[bob1]

You could try gradient centrifugation; I wouldn't trust it to give you a sterile culture at the end though.  

I am presuming you mean that there are no antibiotics rather than antibodies.  Fungizone should still work on the yeast.  However, if you are doing routine culture, I would suggest getting some more cells from a contamination-free source.  I would only treat cells if they are extremely precious.

[Liang]

Liang, on Jun 11 2009, 11:18 AM, said:

I’m looking for a method to separate yeast cells from small human lung tissues. Currently there are no antibodies to this yeast available. I wonder if there are chemical/physical methods to separate yeast cells from human lung tissues infected with this yeast. Thank you in advance.

Sorry I didn't make it clear. Actually my goal is to purify yeast cells from infected patient lung tissue samples. I need to get rid of lung tissues and keep yeast cells for further studies (such as 2-DE analysis). Due to the small quantity of lung samples, gradient centrifugation may not work. Thank you.

[bob1]

OK, in that case, just chuck them into a flask with some medium and the lung cells should die pretty quickly while the yeast should grow.

[gfischer]

bob1, on Jun 15 2009, 07:53 PM, said:

OK, in that case, just chuck them into a flask with some medium and the lung cells should die pretty quickly while the yeast should grow.

I agree.  I was trained as a med tech, and this is how they would get the yeast for isolation in a hospital lab for diagnostic testing.

[Liang]

gfischer, on Jun 16 2009, 06:01 AM, said:

bob1, on Jun 15 2009, 07:53 PM, said:

OK, in that case, just chuck them into a flask with some medium and the lung cells should die pretty quickly while the yeast should grow.

I agree.  I was trained as a med tech, and this is how they would get the yeast for isolation in a hospital lab for diagnostic testing.

Unfortunately currently there is no culture method available for the yeast species I'm working with.

[eldon]

Liang, on Jun 16 2009, 12:00 PM, said:

gfischer, on Jun 16 2009, 06:01 AM, said:

bob1, on Jun 15 2009, 07:53 PM, said:

OK, in that case, just chuck them into a flask with some medium and the lung cells should die pretty quickly while the yeast should grow.

I agree.  I was trained as a med tech, and this is how they would get the yeast for isolation in a hospital lab for diagnostic testing.

Unfortunately currently there is no culture method available for the yeast species I'm working with.

without being able to culture what are you trying to do? eventually culture? present vs absent? cfu? do you have paraffin-embedded lung biopsies?

virulent or do they grow only in an immuno-compromised host?

can you not do qPCR or PCR to detect yeast specific targets? what about probing the cultures with yeast cell surface antigens or simple imaging of the yeast cells...I use a cellometer from Nexcelom...take a picture and adjust parameters to identify your cell type...no dilutions or concentrating required...good for doing counts.

since you cannot culture you must know what you're looking at.. and not trying to identify unknowns as cryptococcus, histoplasma etc.

Edited by eldon, 16 June 2009 - 02:18 PM.

[gebirgsziege]

Maybe you can tell us which yeast you are working with? Maybe someone can give you some hints for culturing.