How To Prepare HBSS Buffer Solution?- A DIY recipe

How prepare HBSS buffer Solution while it is fairly complicated to make which is why a lot of people just buy it. First, you make each solution, the premix, and then the final solution. All solutions should be stored at 4^oC.

simple procedure for the preparation of the HBSS buffer Solution.

1. Prepare 800 mL of distilled water in a suitable container.
2. Add 8 g of Sodium chloride to the solution.
3. Add 0.4 g of Potassium Chloride to the solution.
4. Add 0.14 g of Calcium Chloride to the solution.
5. Add 0.1 g of Magnesium Sulfate Heptahydrate to the solution.
6. Add 0.1 g of Magnesium Chloride Hexahydrate to the solution.
7. Add 0.06 g of Sodium Phosphate Dibasic Dihydrate to the solution.
8. Add 0.06 g of Potassium Phosphate Monobasic to the solution.
9. Add 1 g of D-Glucose (Dextrose) to the solution.
10. Add 0.35 g of Sodium bicarbonate to the solution.
11. Add distilled water until the volume is 1 L.

However, sometimes we need to prepare different solutions, and then we have to mix them before use.

Preparation of the solution for HBSS Buffer:

• Autoclave the distilled water for the preparation of the solution. after autoclaving place allow it to cool down.
• After sterile distilled water is cool down prepare the different stock solutions as mentioned below.

Solution A:

1. Take 90 ml of the distilled water and dissolve the following ingredient.
2. 8.0 g NaCl
3. 0.4 g KCl
4. g glucose
5. Once all the chemicals are dissolved make it up to 100 ml with sterile distilled water.

Solution B:

1. Take 90 ml of the distilled water and dissolve the following ingredient.
2. 0.358 g Na2HPO4 (anhydrous)
3. 0.60 g KH2PO4
4. Once all the chemicals are dissolved make it up to 100 ml with sterile distilled water.

Solution C:

1. Add 0.72 g CaCl2 to 50ml of Sterile distilled water

Solution D:

1. Add 1.23 g MgSO4x7H2O to 50ml of Sterile distilled water

Solution E:

1. Add 0.35 g NaHCO3 to 10ml of distilled H2O

Hank’s Buffered Salt Solution (HBSS) Premix

Combine the solutions in the following order:

• 10 ml Solution A
• 1 ml Solution B
• 1 ml Solution C
• 86 ml distilled H₂O
• 1 ml Solution D

Hank’s Buffered Salt Solution (HBSS) full strength (mix prior to use)

• 9.9 ml Hank’s Premix
• 0.1 ml solution E

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