1X PBS recipe: This guide explains how to prepare phosphate-buffered saline from the required salts, adjust it to pH 7.4 and scale the method safely for routine biological research.
Overview
What is PBS buffer?
Phosphate-buffered saline, usually abbreviated to PBS, is a water-based salt solution widely used in biological research. Its phosphate system helps resist changes in pH, while sodium and potassium salts provide an ionic environment suitable for many routine laboratory procedures.
A commonly used 1X formulation contains 137 mM sodium chloride, 2.7 mM potassium chloride, 10 mM disodium hydrogen phosphate and 1.8 mM monopotassium phosphate. The final pH is commonly adjusted to 7.4, although some protocols use pH 7.2.
PBS is used for washing cells, diluting reagents, preparing samples, immunoassay workflows and other procedures where a simple buffered saline is appropriate. The best formulation depends on the downstream assay, particularly whether calcium, magnesium, detergents or chelating agents are required.
Buffered
Helps maintain pH within a biologically useful range during routine handling.
Versatile
Used across cell biology, immunoassays, microscopy and sample preparation.
Easy to prepare
Can be prepared as a 1X working solution or a concentrated 10X stock.
Composition
PBS buffer composition tables
The formulation below is a widely used PBS recipe for 1 litre of solution. Different commercial and published formulations can vary slightly, so match the recipe to your experimental protocol where required.
Typical PBS pH
Many protocols use PBS at pH 7.4. Some applications specify pH 7.2, so follow the requirements of the assay rather than assuming one formulation is universal.
Recipe scaling
Scale a 1X PBS recipe
For the standard formulation above, scale each reagent in direct proportion to the required final volume.
| Final volume | NaCl | KCl | Na₂HPO₄ | KH₂PO₄ |
|---|---|---|---|---|
| 100 mL | 0.80 g | 0.020 g | 0.144 g | 0.024 g |
| 250 mL | 2.00 g | 0.050 g | 0.360 g | 0.060 g |
| 500 mL | 4.00 g | 0.100 g | 0.720 g | 0.120 g |
| 1 L | 8.00 g | 0.200 g | 1.440 g | 0.240 g |
For 1X PBS prepared from a 10X stock, combine 1 part 10X PBS with 9 parts water.
Preparation
PBS buffer preparation protocol
This protocol describes preparation of 1 litre of the standard 1X formulation shown above.
Add water
Add approximately 800 mL of high-purity water to a clean vessel with sufficient capacity for mixing.
Add the salts
Add 8.0 g NaCl, 0.2 g KCl, 1.44 g Na₂HPO₄ and 0.24 g KH₂PO₄.
Mix until dissolved
Stir until all components are fully dissolved before adjusting pH or final volume.
Check and adjust pH
Measure the pH using a calibrated pH meter. Adjust to the protocol requirement, commonly pH 7.4, using acid or base as appropriate.
Bring to final volume
Add high-purity water to a final volume of 1 litre and mix thoroughly.
Sterilise if required
For applications requiring sterile PBS, use an appropriate validated sterilisation method such as filtration or autoclaving, where compatible with the intended workflow.
Label and store
Record the formulation, concentration, pH, preparation date, preparer and any sterilisation step. Store according to the requirements of your laboratory procedure.
Good laboratory practice
Use calibrated balances and pH meters, record reagent lot numbers where traceability matters, confirm the hydration state of phosphate salts, and avoid using a generic recipe when a validated assay protocol specifies a particular PBS formulation.
Applications
PBS buffer uses in life science research
PBS is useful because it combines simple pH buffering with a saline environment. Its suitability still depends on the assay and on any additives present in the formulation.
Cell biology
Cell and tissue washing
Frequently used to rinse cells or tissues and remove residual medium or unbound material.
Immunoassays
Antibody workflows
Commonly used in washing, dilution and blocking-related workflows, depending on assay chemistry.
Flow cytometry
Cell suspension handling
Can be used as a base buffer for washing or resuspending cells, often with assay-specific additives.
Microscopy
Sample rinsing
Useful for washing fixed or stained samples where the imaging workflow specifies PBS.
Protein research
Reagent and sample dilution
Often used as a simple diluent when phosphate is compatible with the downstream experiment.
Western blotting
PBS and PBST washes
PBS can form the basis of wash buffers. Tween 20 is commonly added when a PBST formulation is required.
Formulation guide
Choosing the right PBS formulation
PBS is not a single universal formulation. Select the buffer according to the chemistry and biology of your experiment.
| Formulation | Key feature | Typical use | Watch for |
|---|---|---|---|
| Standard PBS | Phosphate buffered saline | General washing and dilution | Confirm compatibility with the assay |
| PBS without Ca²⁺/Mg²⁺ | No added divalent cations | Many routine cell handling workflows | Do not substitute when Ca²⁺ or Mg²⁺ is required |
| PBS with Ca²⁺/Mg²⁺ | Contains divalent cations | Protocols where cations support cell or protein interactions | Can be unsuitable where chelation or dissociation is needed |
| PBST | PBS plus non-ionic detergent | Immunoblot and immunoassay washing | Detergent concentration affects stringency |
| PBS plus EDTA | Includes a chelating agent | Selected cell or sample handling workflows | EDTA can interfere with metal-dependent processes |
Procurement
When ready-made PBS can be the better option
Preparing PBS in-house is straightforward, but a ready-made buffer can reduce preparation time and help standardise repeated workflows. The purchasing decision should be based on the experimental specification rather than convenience alone.
1. Match the formulation
Confirm pH, concentration, ions and additives.
2. Review documentation
Check the product specification, SDS and lot documentation where applicable.
3. Check research grade
Select the grade and testing appropriate for the intended research workflow.
4. Consider consistency
For repeated experiments, standardised preparation can reduce avoidable process variation.
Frequently asked questions
PBS buffer FAQs
What is the purpose of PBS buffer?
PBS provides a buffered saline environment for many routine biological procedures, including washing, dilution and sample handling. Whether it is suitable depends on the downstream experiment.
What is the pH of 1X PBS?
A common target is pH 7.4, although some protocols specify pH 7.2. Always use the value required by the method.
How do I make 1X PBS from 10X PBS?
Use a 1:10 dilution. For example, combine 100 mL of 10X PBS with sufficient water to reach a final volume of 1 litre.
Can PBS be sterilised?
Yes. Depending on the formulation and laboratory procedure, PBS can be sterilised by an appropriate method such as filtration or autoclaving. Use a validated method for the intended application.
What is the difference between PBS and PBST?
PBST is PBS containing a surfactant, commonly Tween 20. It is often used for wash steps in immunoassays and western blotting.
Should I use PBS with or without calcium and magnesium?
That depends on the protocol. Divalent cations can support some biological interactions but may be undesirable in dissociation, chelation or other cation-sensitive workflows.
Is every commercial PBS product equivalent?
No. Formulations can differ in phosphate composition, pH, calcium and magnesium content, additives, sterility and grade. Compare specifications before substituting one product for another.
Reference formulation
The 1X and 10X composition values in this guide are based on the widely used Cold Spring Harbor Protocols PBS recipe: 137 mM NaCl, 2.7 mM KCl, 10 mM Na₂HPO₄ and 1.8 mM KH₂PO₄ for 1X PBS. Published and commercial PBS formulations can vary.
ABMIUM research supply
Need a research reagent with clearer specifications?
ABMIUM helps researchers compare research reagents using product documentation, supplier provenance and application-relevant information.
For research use only. This article is for general research information and does not replace a validated laboratory protocol or product-specific instructions.