I used reference wild-type maize (W22 and B73) leaf protoplasts without any fluorescence protein introduced. But I saw some odd autofluorescence particles in some (not all) protoplasts, some cells have one, and some cells have more than one. The figure attached showed two different protoplasts observed through green fluorescence (left column) and UV (right column) filter.
Have you even seen these particles before? What are they? Is there any way to get rid of them? Thanks for your answer!!
Hi, Zhiyong, did you mean "plastids"? do you have any evidence to support? Why can they generate green fluorescence?
If I transiently transfect a GFP construct into the proroplasts and I want to sort the ones with the expressed GFP, those protoplasts with the endogenous autofluorescence particle will mess up my result. Do you have any idea to solve this problem? Thanks.
they are debris. this is common in protoplast preps. they will auto florescent in any spectra, mostly likely clumps of cell walls or other particles.
normally you can find a field of view lack of those debirs. also if you try to focus the protoplasts the debris are usually floating and out of focus sometimes. if you really need to get rid of them completely you can try to centrifuge your cells with low speed (typically
- Badges
- Science topic
- Similar topics
- Analytical Chemistry
- Column