Optimization of preservation conditions for the detection of enolase from midgut tissues of field collected aedes albopictus / Wan Muhammad Hanif Husin

Aedes albopictus is a vector responsible for dengue outbreaks. The etiological agent, the dengue virus (DENV) must cross the vector's midgut epithelial cells to establish infection. In midgut tissues, enolase appears to be an important protein receptor which takes part in the entrance of DENV that can be detected by observing the protein profiles in sodium dodecylsulphate polyacrylamide gel electrophoresis (SDS-PAGE). Field collected samples are precious yet very fragile and can be easily degraded if not collected, handled or preserved properly. Important protein markers can remain undetected leading to false negative results. To date, the knowledge on the optimum preservation conditions of midgut proteins is currently limited, which forms the rationale of this study. Samples of Ae. albopictus collected from Kolam Basah 8, UiTM Puncak Alam were reared until adulthood under insectary conditions and dissected to obtain midgut tissues. The harvested tissues were then preserved in three medium namely; phosphate buffered saline (PBS), PBS supplemented with anti-protease cocktail (PBSpi), and cell lysis buffer (CLB) at 20°C, 4°C, and room temperature (RT) for 3 days. SDS-PAGE was performed after 3 days of preservation and the result revealed the protein of interest (Mr 57 kDa) at -20°C and 4°C when preserved in PBS, PBSpi, and CLB. However, the protein was not detected at RT in all 3 media. Findings of this study indicates that midgut tissues from field collected Ae. albopictus can be well preserved upon storage in PBS, PBSpi, and CLB media at laboratory conditions of -20°C and 4° for the purpose of enolase detection. The identity of enolase should be further confirmed in forthcoming studies using mass spectrophotometry, amino acid sequencing or Western Blot.