Microalgae cells have the potential to rapidly accumulate lipids, such as triacylglycerides that contain fatty acids important for high value fatty acids (e.g., EPA and DHA) and/or biodiesel production. However, lipid extraction methods for microalgae cells are not well established, and there is currently no standard extraction method for the determination of the fatty acid content of microalgae. This has caused a few problems in microlagal biofuel research due to the bias derived from different extraction methods. Therefore, this study used several extraction methods for fatty acid analysis on marine microalga Tetraselmis sp. M8, aiming to assess the potential impact of different extractions on current microalgal lipid research. These methods included classical Bligh & Dyer lipid extraction, two other chemical extractions using different solvents and sonication, direct saponification and supercritical CO₂ extraction. Soxhlet-based extraction was used to weigh out the importance of solvent polarity in the algal oil extraction. Coupled with GC/MS, a Thermogravimetric Analyser was used to improve the quantification of microalgal lipid extractions. Among these extractions, significant differences were observed in both, extract yield and fatty acid composition. The supercritical extraction technique stood out most for effective extraction of microalgal lipids, especially for long chain unsaturated fatty acids. The results highlight the necessity for comparative analyses of microalgae fatty acids and careful choice and validation of analytical methodology in microalgal lipid research.