Hello group,
Has any one attempted to use coverslip thickness glass slides rather than standard glass slides for attaching formalin fixed paraffin embedded tissues slices obtained after slicing a paraffin block?
Following immunostaining my goal is to look at sub-cellular (mitochondria) structures and with the current approach of putting tissues on glass slides and imaging using a 63X objective on a confocal all I see is a smear of stain in the cytoplasm. I feel that putting tissues on a coverslip thickness glass slide will the best option but very tricky also. This brings to series of questions
1) Are there special charged coverslip thickness slides available for this purpose so that the tissue sticks properly?
2) Is there an insert available in which the coverslip thickness slide could be slipped so that it won't break during successive handling steps.
3) What is your preferred aqueous mounting medium that would prevent photobleaching?
I will be happy to hear your thoughts on my relevant points or if you have a methodology for imaging sub-cellular structures.
Best,
Gaurav.
Dear Gaurav,
Based on my knowledge there is no differences between the coverslips thickness except than the length of them, usually applying extra mounting media would cause to bad quality images with high background. what usually I do is that I put a very small drop of mounting media in the bottom of slide and then place the coverslip from the same side of glue and t let it to move toward the section, also I keep the slides there for half an hour without moving anywhere. If you want to see them in high magnification oil will be very helpful and most of the time you can find it in microscopy room.
To seen structures in sub cellular level (mitochondria) electron microscoe also is good choice unless you are looking for specific markers.
Hope this can help.
Good Luck
Masoumeh
Dear Gaurav,
as you use confocal microscopy, you can do optical sections and you should be able to get perfect pictures from your immunostained FFPE material on normal glass slides. To immunostain tissue collected on coverslips did not work in our hands (sections did not stick well, bigger coverslips brake easily, are difficult to view under the microscope.
Regards, Fred
- Badges
- Science topic
- Similar topics
- Physical Sciences
- Materials Science
- Materials
- Inorganic Materials
- Ceramics
- Glass