Using Phosphate-Buffered Saline (PBS) in Biochemical and Cell Biology Research

Biochemistry, Molecular Biology, and Cell Biology Protocols >> Using Phosphate-Buffered Saline (PBS) in Biochemical and Cell Biology Research

PBS is often used in cell biology experiments to maintain the osmolarity of the cells. It contains salt ions, which balances the amount of salt ions inside the cell. If the cells are immersed into a solution that has too many salt ions, water will leak out from the cell, causing the cell to shrink. Conversely, if the cells are immersed into a solution that has too few salt ions, water will enter the cell, causing the cell to burst. Therefore, it is critical when performing cell biology experiments to maintain the cells at a certain osmolarity. PBS is at the correct osmolarity to keep the cells in an isotonic state.

PBS is also often used as a buffer in biochemistry experiments to maintain the pH of proteins. It can be used to dissolve peptide or protein samples directly, and to store the protein or peptide in that solution. Proteins require a certain pH range to maintain neutrality or charges on certain amino acids, which allow the structure of the protein to be maintained in its native state. Otherwise, the structure of the protein may become denatured. Besides using PBS to directly dissolve proteins, PBS is also highly useful as a buffer to maintain the pH in biochemical assays such as Western blots.

One note about PBS is that it should not be combined in a solution containing calcium, as calcium and phosphate can result in precipitate formation. Instead, a different buffer such as Tris or HEPES should be applied. If performing cell biology experiments, the salt concentration should still be maintained to maintain the isotonicity of the cells.

Making PBS

You can either make a 10x solution from below and dilute to 1x with double distilled water:

10x Phosphate Buffered Saline—PBS
NaCl            1.37 M
KCl              27mM
Na₂HPO₄80mM
KH₂PO₄     20mM
double distilled water to 1L or other desired volume
stir well
filter if desired
store at room temperature

pH should already be at pH 7.4; just remember to measure the pH of the 1x solution and not the 10x solution

You can also make a 1x PBS solution directly:

1x Phosphate Buffered Saline—PBS
NaCl            137 mM
KCl                  2.7mM
Na₂HPO₄8   mM
KH₂PO₄          2 mM
water to 1L or other desired volume
stir well
filter if desired
can store at room temperature