Product Class: Other

NEBExpress® E. coli Lysis Reagent
25


Catalog #P8116

Product Introduction

NEBExpress E. coli Lysis Reagent is a chemical lysis solution composed of a proprietary mix of non-ionic and zwitterionic detergents and Tris-based buffer. It allows disruption of E. coli cells without denaturing soluble proteins.

  • Ready-to-use liquid that is stable at room temperature
  • Gentle yet highly active formulation of detergents in Tris buffer
  • Scalable lysis reactions from small to large bacterial cell pellets and compatible with high throughput workflows 
  • Suitable for extracting proteins, especially thermosensitive proteins vulnerable to mechanical lysis procedures
  • Compatible with analyses such as SDS-PAGE, Western blots, activity assay, immunoprecipitation, and downstream purification 
  • Suitable for disrupting most Gram-negative bacterial cells
  • Lysis efficiency increases 2-fold when used in combination with NEBExpress T4 Lysozyme (NEB# P8115)
 

Product Information

Description

NEBExpress® E. coli Lysis Reagent is a chemical lysis solution composed of a proprietary mix of non-ionic and zwitterionic detergents and Tris-based buffer. It allows disruption of E. coli cells without denaturing soluble proteins.

Figure 1: Optimal protein extraction with NEBExpress T4 Lysozyme in the presence of NEBExpress E. coli Lysis Reagent.

P8115 Figure 2

T7 Express E. coli  expressing vGFP was lysed with NEBExpress T4 Lysozyme (T4L) in 50 mM Tris-HCl pH 7.5 or in NEBExpress E. coli Lysis Reagent (NEB #P8116S): 4 UOD600 of cell pellet were resuspended in 200 µL of Tris buffer or NEBExpress E. coli Lysis Reagent and lysed for 5 minutes at room temperature in the presence of NEBExpress T4 Lysozyme at 0, 0.1 or 1 ug per 1 mL of cell suspension. The soluble proteins were harvested by centrifugation and analyzed on SDS-PAGE.

This product is related to the following categories:
Competent Cells
This product can be used in the following applications:
High-throughput cloning and automation solutions,
Protein Expression in E. Coli,
Protein Expression

Properties & Usage

Features

  • Scalable lysis reactions from small to large bacterial cell pellets and compatible with high throughput workflows
  • Suitable for extracting proteins, especially thermosensitive proteins vulnerable to mechanical lysis procedures
  • Fast and simple protocol: harvested cells are resuspended and briefly incubated in the Lysis Reagent to extract soluble proteins. Insoluble cell debris is removed by centrifugation, and the clarified extract is ready to use and compatible with affinity resins.
  • Compatible with analyses such as SDS-PAGE, Western blots, activity assays, immunoprecipitation, and downstream purification 
  • Suitable for disrupting most Gram-negative bacterial cells
  • Small (100 ml) and large kits (5 x 100 ml) are suitable for lysing 20 to 100 grams of cells, respectively.
 

Product Notes

  1. NEBExpress E. coli Lysis Reagent is compatible with: 

    • Tris, phosphate, and PIPPS-based buffers ranging from pH 5.5-10.
    • EDTA. To prevent interference with metal affinity resins use < 0.1-0.5 mM EDTA. If using NEBExpress Ni Resin (NEB #S1428S) or Ni Spin Columns (NEB #S1427S/L), there is no limitation on EDTA use, as the resin is chemically resistant to EDTA chelation.
    • < 1 M NaCl (precipitation will occur at or above 1 M NaCl).
    • Protease inhibitors 
    • Reducing agent such as DTT, β-mercaptoethanol, TCEP, THP. To prevent interference with metal affinity resins, use 1-2 mM β-mercaptoethanol or 1-5 mM TCEP. NEBExpress Ni Resin (NEB #S1428) and Ni Spin Columns (NEB #S1427S/L) are both chemically resistant to these reducing agents. 
    • BCA Protein assay
    • SDS-PAGE and Western blots
    • 1 µL NEBExpress® T4 Lysozyme (NEB# P8115) per 1 mL of lysate for improved protein extraction efficiency. The lysate may become viscous due to the release of DNA. For optimal lysis yield, add 1mM CaCl2 and 1 µL of Micrococcal Nuclease (NEB #M0247) per 1 mL of lysate, mix and incubate for 5 minutes at room temperature.
     
  2. NEBExpress E. coli Lysis Reagent can be added directly to frozen cell pellets; however, allow this mixture to reach room temperature as the lysis will be more efficient.
  3. Use 0.025 – 0.075 mL NEBExpress E. coli Lysis Reagent per unit OD600 (UOD600) cells harvested. For example, for a 5 mL culture harvested at OD600 of 1, 5 UOD600 of cells are obtained (5 mL x 1 OD600 = 5 UOD600), thus requiring 0.125 to 0.375 mL of Lysis Reagent. 
  4. If weighing the cell pellet, use 5 mL NEBExpress E. coli Lysis Reagent per 1 gram of cell paste. Note that cells resuspended in Lysis Reagent at higher cell densities will not lyse as efficiently.
  5. Extraction efficiency is strain dependent. Efficiency is highest with BL21 strains, its derivatives with DE3 or lysY, E. coli K-12 strains, and various other Gram-negative bacteria (e.g., Salmonella, Vibrio, Pseudomonas).
  6. Storing NEBExpress E. coli Lysis Reagent at 4°C or below may cause precipitation of the detergents. However, the lysis efficiency of the reagent can be fully restored by incubation at room temperature with gentle mixing to dissolve any precipitate.

Protocols, Manuals & Usage

Protocols

  1. Protocol for cell lysis using NEBExpress E. coli Lysis Reagent (NEB #P8116)

FAQs & Troubleshooting

FAQs

  1. How much NEBExpress® E. coli Lysis Reagent should I use?
  2. Why do I see precipitation in the NEBExpress® E. coli Lysis Reagent?
  3. Can I use NEBExpress® E. coli Lysis Reagent that has been frozen?
  4. Should I chill the NEBExpress® E. coli Lysis Reagent prior to use?
  5. Should I freeze the cell pellets prior to lysing with NEBExpress® E. coli Lysis Reagent?
  6. What buffer is present in the NEBExpress® E. coli Lysis Reagent?
  7. Can I supplement the NEBExpress® E. coli Lysis Reagent with NaCl?
  8. Can I change the pH of NEBExpress® E. coli Lysis Reagent?
  9. Is the NEBExpress® E. coli Lysis Reagent compatible with EDTA?
  10. Can I add a protease inhibitor to the NEBExpress® E. coli Lysis Reagent reaction?
  11. Do I need to add lysozyme to aid in lysis?
  12. My lysate is viscous; how can I resolve this?
  13. Can I use a reducing agent with NEBExpress® E. coli Lysis Reagent?
  14. Does NEBExpress® E. coli Lysis Reagent lyse gram-positive bacterial cells or yeast cells?
  15. Does NEBExpress® E. coli Lysis Reagent lyse gram-negative bacterial cells?
  16. My protein forms inclusion bodies. Can I still use this reagent to lyse my cells?
  17. Can I add NEBExpress® E. coli Lysis Reagent directly to my cell culture to lyse the cells?

Troubleshooting

Quality, Safety & Legal

Quality Assurance Statement

Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.

Specifications

The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

Certificate Of Analysis

The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

Safety DataSheets

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

Legal and Disclaimers

Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email busdev@neb.com.

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.

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