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AAT Bioquest

What criteria should I use to determine the type of lysis buffer required for an experiment?

Posted April 26, 2023


Answer

Using the right type of lysis buffer is critical to obtaining accurate results. The right buffer helps provide vital pH control and solution stability without interfering with the ongoing reaction. 

These are some of the criteria you should use to determine the type of lysis buffer required for an experiment: 

  • pH range – A pH range or pKa of 6.0 to 8.5 is ideal for most experiments as this is the optimal pH range for most biological processes. 
  • Inertness – The buffer you choose must not participate in, influence, or interfere with any of the components, reactions, or biological processes in your experiment. 
  • Minimal effect on salts – The components of the buffer should not, in any way, interfere or interact with ions involved in the biochemical reaction you’re studying.  
  • Good solubility in water – Buffers that exhibit high water solubility are preferable as biological processes favor aqueous environments. 
  • Chemical stability – A buffer that is chemically stable will resist getting oxidized or degraded in any way under working conditions.  
  • Resistant to changes in concentration, ionic composition and temperature – The buffering capabilities of your chosen buffer should not be affected by the concentration, ionic strength, and temperature of the medium. Making the buffer at the temperature you’ll be using it at will give you the most accurate results. 
  • Membrane-impermeant – The chosen buffer must not interfere with or penetrate the cell wall so it does not get into the cytosol. 
  • Non-toxic to cells – The lysis buffer you choose should not kill your sample. 
  • Minimum optical absorbance – Choosing a buffer that only absorbs light at wavelengths < 230 nm will help to prevent interferences in spectrophotometric assays.
  • Known complex-forming tendency with metal ions – Choose a buffer with low metal-binding constant for enzyme assays such as PCR. If using a metal is required for your assay, choose a buffer that will not form a complex with that metal. 
  • Convenient and cost-effective – Buffer preparation and purification should meet all the above criteria and should also be easy and inexpensive.
Additional resources

Solubilization of proteins: the importance of lysis buffer choice

Buffers and Lab Consumables

ReadiUse™ Viral RNA Lysis Buffer