Comparison of F-actin fluorescent labeling methods in pollen tubes of Lilium davidii.
Fluorescence labeling of F-actin in pollen tubes by various methods has produced inconsistent results in the literature. Here, we report that EGTA, which was always used in fixative buffers in the past and thought to help cytoskeleton stabilization, can significantly affect F-actin distribution and lead to the formation of thick F-actin bundles at the tip of the pollen tube. We also found that vacuum-infiltration for the first 5 min during pollen tube fixation can better preserve normal cytoplasm structure and F-actin distribution. In contrast, m-maleimidobenzoic acid N-hydroxysuccinimide ester (MBS) treatment before chemical fixation resulted in a shortening of the free zone of thick F-actin bundles in the pollen tube tip. Taken together, our results suggest that exclusion of EGTA and MBS from the fixative buffer, in combination with vacuum-infiltration in the first 5 min of fixation, can improve F-actin fluorescence labeling in pollen tubes of Lilium davidii.
Wang
Li
L
State Key Laboratory of Plant Physiology and Biochemistry, College of Biological Sciences, China Agricultural University, Beijing, 100094, PR China.
Liu
Yi-Min
YM
Li
Yan
Y
D000199:Actins / Q000378:metabolism*; D002021:Buffers; D004533:Egtazic Acid / Q000737:chemistry; D035264:Flowers / Q000166:cytology / Q000378:metabolism*; D005456:Fluorescent Dyes; D027762:Lilium / Q000166:cytology / Q000378:metabolism*; D018613:Microscopy, Confocal; D013388:Succinimides / Q000737:chemistry; D016707:Tissue Fixation
D000199:Actins; D002021:Buffers; D005456:Fluorescent Dyes; D013388:Succinimides; C072252:m-maleimidobenzoyl-N-hydroxysulfosuccinimide ester; D004533:Egtazic Acid