Common Lysis Buffers

VIEW LYSIS BUFFERS

Lysis buffers are specialized solutions used to break open (lyse) cell membranes and release intracellular contents, such as DNA, RNA, proteins, lipids, or organelles, for downstream experiments including:

Choosing the right buffer is crucial: it can affect yield, integrity, specificity, and the success of downstream analyses.

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What is a Lysis Buffer?

A lysis buffer is a chemical solution that disrupts cell membranes (or walls) to liberate internal components. These reagents contain different chemicals specialized for degrading certain membranes. It typically includes a buffer (to stabilize pH), salts, detergents, inhibitors, and chelators.

Mechanism-of-cellular-Lysis

Image 1: The mechanism of cellular lysis. Using a detergent within a lysis buffer solution, lysates may be extracted from specific cellular compartments for later analysis.

How to Choose a Lysis Buffer: Step-by-Step Guide

  1. Define the analyte(s) of interest
    • Are you after soluble cytoplasmic proteins, nuclear proteins, membrane proteins, mitochondrial proteins, or RNA/DNA?
    • Tough membranes (plant, yeast, bacteria) may need more aggressive detergents or physical lysis.
  2. Understand the downstream applications
    • For enzymatic assays, you need buffers that preserve activity (i.e. mild, non-denaturing).
    • In Western blot, immunoprecipitation, or ELISA, protein integrity and solubility matter.
    • In mass spectrometry, low detergent concentration or detergent-free may be preferred.
  3. Balance strength vs. specificity
    • Strong buffers (e.g. RIPA) extract more, but risk extracting unwanted proteins or destroying protein complexes.
    • Mild buffers preserve complexes but may give lower yield or miss membrane/nuclear proteins.
  4. Consider additives
  5. Check compatibility
    • Detergents can interfere with downstream assays (e.g., detergents may hamper mass spec, enzyme kinetics, or membrane protein reconstitution).
    • Buffer pH and salt concentrations should be compatible with downstream enzymatic reactions or antibody binding.
  6. Pilot test / optimization
    • Test 2-3 buffer types in small scale to compare yield, specificity, and downstream performance.
    • Adjust detergent concentration, incubation time, temperature as needed.

Common Lysis Buffers & Applications

Key Features  

Ionic + non-ionic detergents (e.g. SDS / deoxycholate + NP-40/Triton-X), salts, buffer 

Cell Location Targeted 

Cytoplasmic, nuclear, membrane, mitochondrial (broad spectrum) 

Applications 

Western blot (whole cell extracts), immunoprecipitation, co-IP, proteomics 

Considerations/ Limitations 

Strong solubilization, but may disrupt protein-protein interactions; SDS can interfere with enzyme assays unless removed 

 

Browse our selection of RIPA Lysis Buffers here 

Key Features  

NP-40 detergent, mild salts, buffering agents 

Cell Location Targeted 

Cytoplasmic, membrane (soluble proteins, membrane-associated) 

Applications 

Preserving protein complexes, co-IP, Western blot, enzyme assays, ELISA 

Considerations/ Limitations 

Gentle; lower yield for nuclear or tightly membrane-bound proteins 

 

Browse our selection of NP-40 Lysis Buffers here 

Key Features  

Non-ionic detergent Triton X-100 with Tris or HEPES buffer 

Cell Location Targeted 

Cytoplasmic, membrane-associated proteins, cytoskeletal proteins 

Applications 

Membrane protein studies, Western blot, immunofluorescence, co-IP 

Considerations/ Limitations 

Mild, but residual detergent may affect downstream assays; may under-lyse nuclei 

Browse our selection of Triton X-100 Lysis Buffers here 

Key Features  

Zwitterionic detergent (CHAPS), salts, buffer 

Cell Location Targeted 

Cytoplasmic, membrane, soluble nuclear proteins (good balance) 

Applications 

Co-IP, native PAGE, Western blot, protein–protein interaction studies 

Considerations/ Limitations 

Preserves activity/structure; may not solubilize very hydrophobic proteins; higher cost

 

Browse our selection of CHAPS Lysis Buffers here 

Key Features / Detergents & Components 

Chaotropic agents, detergents, RNase inhibitors 

Cell Location Targeted 

RNA (cytoplasmic + nuclear) 

Applications 

RNA-seq, RT-PCR, transcriptomics, RNA stabilization 

Considerations/ Limitations 

Inactivates RNases fast; proteins usually denatured — not suitable for protein assays 

 

Browse our selection of RNA Lysis Buffers here 

Key Features  

Hypotonic buffer or selective lysis buffer 

Cell Location Targeted 

Red blood cells (targets hemoglobin-containing cells; preserves WBCs) 

Applications 

Flow cytometry, immunophenotyping, DNA/RNA/protein extraction from WBCs 

Considerations/ Limitations 

Over-lysis can harm non-RBC cells; timing critical 

 

Browse our selection of RBC Lysis Buffers here 

Practical Tips & Best Practices

  • Use inhibitors: Add protease and phosphatase inhibitors stock should be added just before lysis. Some degrade over time.
  • Keep everything cold (on ice) during lysis and centrifugation to reduce protease activity.
  • Work fast: Minimize the time between cell harvest and protein/RNA stabilization.
  • Clarify lysates by centrifugation to remove debris; this reduces background in downstream analysis.
  • Consider detergent removal when needed: Some assays require detergent-free samples; use dialysis, spin filters, or detergent-removing resins.
  • Validate lysis efficiency: Use controls like Western blots for cytosolic vs nuclear markers or microscopy to confirm lysis.

Troubleshooting common issues:

Problem

Possible Cause

Suggested Fixes
Low Yield Weak buffer / low detergent / insufficient incubation time Use stronger buffer (e.g. RIPA), increase incubation, increase detergent concentration
Loss of protein activity Harsh detergent / proteases / inappropriate pH Use mild buffer, include inhibitors, optimize pH, use reducing agents only if needed and if disulfide protein linkage is not critical.
High background in assays Debris not cleared / non-specific proteins pulled down Centrifuge well; preclear; use more specific buffer; adjust detergent types
Membrane proteins insoluble Wrong type of detergent; too mild Use detergents better at solubilizing membranes (ionic or stronger non-ionic or zwitterionic)

Boston BioProducts’ Lysis Buffer Options & Custom Formulation

  • Browse our catalog of pre-formulated lysis buffers (NP-40, RIPA, CHAPS, Triton X-100, etc.), each optimized for different cell types and analyte locations.
  • Use our Custom Manufacturing / Reagent Builder to design a buffer tailored to your specific needs (detergent choice, inhibitor mix, buffer strength).

Frequently Asked Questions (FAQ)

RIPA is stronger: it uses both ionic and non-ionic detergents and extracts proteins from many compartments (cytosolic, membrane, nuclear, mitochondrial). NP-40 is milder and tends to preserve more native protein interactions, making it better when you want less disruption.

Yes, if you care about preserving protein integrity and post-translational modifications. Proteases can degrade your target proteins, phosphatases can remove phosphorylation. For non-enzymatic readouts (e.g. total protein quantification), inhibitors are less critical.

They can. Some detergents ionize poorly or suppress signals in MS. In these cases, using detergent-free buffers or detergents that are MS-compatible, and removing detergents before analysis (via cleanup or membrane filters) is recommended.

You will often need both chemical and physical methods: stronger detergents, enzymatic treatment (lysozyme, zymolyase), mechanical disruption (bead beating, sonication) or high osmotic pressure. Also ensure sufficient incubation time, temperature, and possibly adding EDTA to help with cell wall weakening.

Keep lysates on ice for short term (hours). For longer storage, freeze at -80 °C in aliquots to avoid repeated freeze-thaw. Add glycerol or stabilizing agents if needed.