Human Tubal Fluid Medium (HTF)
| Reagent | Sigma-Aldrich catalog no. | Molecular weight | Final concentration (mm) | Final concentration (g/L) |
|---|---|---|---|---|
| NaCl | S5886 | 58.44 | 101.6 | 5.9375 |
| KCl | P5405 | 74.55 | 4.69 | 0.3496 |
| MgSO4 · 7H2O | M7774 | 246.5 | 0.20 | 0.0493 |
| KH2PO4 | P5655 | 136.09 | 0.37 | 0.0504 |
| CaCl2 · 2H2O | C7902 | 147 | 2.04 | 0.3 |
| NaHCO3 | S5761 | 84.01 | 25.0 | 2.1 |
| Glucose | G6152 | 180.16 | 2.78 | 0.5 |
| Na-pyruvate | P4562 | 110.0 | 0.33 | 0.0365 |
| Na-lactate 60% syrup | L7900 | 112.1 | 21.4 | 2.39893.998 g (3.42 mL) |
| Penicillin-G | P4687 | 372.5 | 0.075 | |
| Streptomycin sulfate | S1277 | 1457.4 | 0.05 | |
| Phenol Red (5%) | P0290 | 0.20 mL | ||
| BSA (embryo tested) | e.g., Equitech-Bio BAC62-0050 (or similar) | 4.0 |
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1. Add each chemical in order and mix carefully. Note that 60% syrup of sodium lactate has a large osmotic effect and must be measured very accurately.
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2. Dissolve CaCl2 in 5 mL of H2O separately and add it to the solution.
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3. Before adding BSA, apply gas for 5 min with 5% CO2, 5% O2 balanced with N2, using a 1-mL pipette submerged in the medium or place in the incubator.
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4. Add BSA after gassing, and incubate to dissolve.
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5. Filter through a 0.22-µm filter.
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6. Measure osmolarity; it should be 290% ± 5% Osm.
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7. Gas before refrigerating.
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8. Store no longer than 2 wk at 4°C. Can be kept frozen for 6 mo or more depending on the temperature.
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9. Always gas medium with CO2 after every use. The medium should be salmon pink at the time of use.
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See Quinn et al. (1985).
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