title: Imaging the Drosophila retina: zwitterionic buffers PIPES and HEPES induce morphological artifacts in tissue fixation.
abstract: BACKGROUND
  Tissue fixation is crucial for preserving the morphology of biological structures and cytological details to prevent postmortem degradation and autolysis. Improper fixation conditions could lead to artifacts and thus incorrect conclusions in immunofluorescence or histology experiments. To resolve reported structural anomalies with respect to Drosophila photoreceptor cell organization we developed and utilized a combination of live imaging and fixed samples to investigate the exact biogenesis and to identify the underlying source for the reported discrepancies in structure.
  
  
  RESULTS
  We found that piperazine-N,N'-bis(ethanesulfonic acid) (PIPES) and 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), two zwitterionic buffers commonly used in tissue fixation, can cause severe lumen and cell morphological defects in Drosophila pupal and adult retina; the inter-rhabdomeral lumen becomes dilated and the photoreceptor cells are significantly reduced in size. Correspondingly, the localization pattern of Eyes shut (EYS), a luminal protein, is severely altered. In contrast, tissues fixed in the phosphate buffered saline (PBS) buffer results in lumen and cell morphologies that are consistent with live imaging.
  
  
  CONCLUSIONS
  We suggest that PIPES and HEPES buffers should be utilized with caution for fixation when examining the interplay between cells and their extracellular environment, especially in Drosophila pupal and adult retina research.
authors: Department of Biology, Indiana University, 1001 East Third St, Bloomington, IN, 47405, USA. jingnie@umail.iu.edu.|Department of Otolaryngology-Head and Neck Surgery, Indiana University School of Medicine, Indianapolis, IN, 46202, USA. jingnie@umail.iu.edu.
authors: Jing
authors: J
authors: Nie
authors: Department of Biology, Indiana University, 1001 East Third St, Bloomington, IN, 47405, USA. smahato@indiana.edu.
authors: Simpla
authors: S
authors: Mahato
authors: Department of Biology, Indiana University, 1001 East Third St, Bloomington, IN, 47405, USA. azelhof@indiana.edu.
authors: Andrew C
authors: AC
authors: Zelhof
mesh_terms: D017738:Alkanesulfonic Acids; D000818:Animals; D030801:Animals, Genetically Modified; D016477:Artifacts; D002021:Buffers; D004330:Drosophila; D006531:HEPES; D010879:Piperazines; D012160:Retina / Q000033:anatomy & histology*; D016707:Tissue Fixation
chemical_list: D017738:Alkanesulfonic Acids; D002021:Buffers; D010879:Piperazines; C008916:piperazine-N,N'-bis(2-ethanesulfonic acid); D006531:HEPES
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