TC, TG experimental method - Huaqiang Electronic Network
First, the tissue sample preparation.
1
Use an isotonic buffer solution, by weight.
1
:
9
, prepare a homogenate, then centrifuge at 2500 rpm for 10 minutes and collect the supernatant.
10%
Homogenate
TG
,
TC
Determination (extraction is not complete, but can also be measured)
2
Extract using an organic solvent. a: Use anhydrous ethanol as the homogenization medium, prepare the homogenate as described above, and the extraction efficiency is better than that of isotonic buffer. b: Extraction with special organic solvents (such as methanol mixtures) is effective, but due to high volatility, measurement accuracy may be affected. Therefore, it is generally recommended to dry the extract under nitrogen gas and then reconstitute it in absolute ethanol before measurement.
Second, cell culture.
1
, wash the cells with PBS.
2
Add 500 μl of ethanol to the culture dish, allowing it to spread evenly. When the cells appear white, tilt the dish and scrape the cells off using a cell scraper, so they are suspended in ethanol.
3
Transfer the cell-ethanol mixture to a glass homogenizer using a pipette and grind for 3 minutes to form a suspension.
4
, collect the suspension, centrifuge at 4000 rpm for 10 minutes, and take the supernatant.
50 μl
for determination.
Third, milk sample processing.
For fresh or frozen milk samples, extract using absolute ethanol in different ratios (milk sample : absolute ethanol = 1 : 1, 1 : 4, 1 : 9, 1 : 19, 1 : 49, 1 : 99). Vortex thoroughly for 2 minutes, then centrifuge at 4000 rpm for 10 minutes. Collect the clear supernatant and proceed with further analysis.
Based on the dilution factor and the measured results, plot a standard curve and select the linear segment for accurate concentration calculation.
Throughout the process, it's important to maintain consistent conditions and ensure all steps are performed carefully to avoid contamination or inaccurate measurements. Proper documentation of each step is also crucial for reproducibility and data integrity.
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