EDAC fixation increases the demonstrability of biogenic amines in the unicellular Tetrahymena: A flow cytometric and confocal microscopic comparative analysis

Abstract

Earlier experiments demonstrated the presence of hormones of the higher ranked animals in Tetrahymena. In the present experiments two fixatives, paraformaldehyde, which is commonly used and a carbodiimide, EDAC that was recommended by Panula et al. for the immunocytochemistry of histamine, are compared in Tetrahymena for the demonstration of histamine and serotonin by using flow cytometry and confocal microscopy. Both hormone levels were significantly higher after EDAC fixation; serotonin almost doubled and histamine was more than fivefold. The confocal microscopic pictures were clearer and the hormones' localization was easier. The results support earlier observations on the presence of these hormones in Tetrahymena and points to the advantage of EDAC fixation for demonstrating these hormones immunocytochemically.

Optimized Protocol for Retinal Wholemount Preparation for Imaging and Immunohistochemistry

“…Shorter fixation times result in poor tissue preservation but bright punctate labeling. Similar results have been obtained for many other synaptic structural proteins and ion channels by different groups 1–7 . Using the membrane for structural support enables the use of carbodiimide to simultaneously achieve short fixation times, well-defined bright labeling of synaptic structures and adequate overall tissue quality for both wholemount and cryostat section preparations.…”

“…Additionally, there are certain methods of tissue fixation that can be advantageous for immunostaining of neuronal structures, but result in compromised structural stability, precluding their use. A particular example of this is carbodiimide-based fixation, which is superior for staining receptors and hormones 1–7 but is commonly avoided due to the instability of the fixed tissue.…”

Bioactive albumin functionalized polylactic acid membranes for improved biocompatibility

“…EDAC is a well-known heterobifunctional cross-linker used in peptide synthesis; cross-linking proteins to nucleic acids; and preparation of immunoconjugates [42][43][44]. The increase in the total antioxidant activity (amount of quenched TMAMQ) further confirmed that indeed albumin was successfully grafted onto the PLA membranes and that heating albumin helped expose amino acids groups with antioxidant properties buried inside the 3-D structure.…”

The hormonal system of the unicellular Tetrahymena: A review with evolutionary aspects

“…The presence of serotonin (5-hydroxytryptamine, 5HT) was demonstrated as early, as 1966 [6] in Tetrahymena. The presence of the hormone was biochemically as well as immunocytochemically justified [7,8]. The serotonin metabolite 5-hydroxyindole acetic acid was also found and it was supposed that a serotonergic system is functioning in Tetrahymena [7], and serotonin is a chemical mediator [9].…”