Thesis

Investigation of formaldehyde effects on electrophoresis banding patterns: are the effects reversible?

Formaldehyde has long been a commercially available fixative of common use. Formalin is the term used for an aqueous solution of formaldehyde and has been used over decades as a fixative for preserving tissue integrity. Due to formaldehyde cross-linking of proteins molecular analysis of tissues treated with formalin has not been simple. The mechanisms of formalin fixation and reversal are poorly understood. Previous studies in the laboratory have used formalin to preserve sea urchin embryos for cell adhesion studies. A goal of the research in this laboratory is to learn if formaldehyde fixation allows cell surface components in sea urchin embryo structure to retain function. A first step is this project where I investigate how formaldehyde fixation influences bands on polyacrylamide gels of sea urchin embryo material and the standard protein bovine serum albumin. In about a total of three hundred and thirty seven experiments using numerous protocols this project shows that formaldehyde fixation causes bands to disappear or lighten considerably. Before carefully controlling protein concentrations on gels many experiments were prepared with one or more errors occurring. Finally it was found that attention to solubilization protocols and protein quantity standardization provided reliable results. While the effects of formaldehyde on the appearance of bands could not entirely be reversed, a substantial amount of insight was achieved into the methods that may eventually lead to complete formaldehyde crosslink reversal in these model systems.

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