Cell and tissue lysis hub: Difference between revisions

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[[Image:Hub icon.png|right]]
[[Image:Hub icon.png|right]]


This page should point you to the many different general and lab-specific protocols describing tissue and cell lysis and serve as forum for comparison.
This page should point you to the many different general and lab-specific protocols describing tissue and cell lysis and serve as forum for comparison. The ups and downs of various methods from sonication, homogenization, freeze-thaw cycles, and detergent-based lysis are examined below.


== Comparison of lysis methods ==
== Comparison of lysis methods ==
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* problem: heat build up which can denature proteins (proportionate to length of sonication)
* problem: heat build up which can denature proteins (proportionate to length of sonication)
* precaution: do on ice and sonicate intermittantly
* precaution: do on ice and sonicate intermittantly
material: ultrasonication bath or rods


=== Homogenisation ===  
=== Homogenisation ===  
* best for animal tissue; less suitable for cells
* best for animal tissue; less suitable for cells
* precaution: do on ice to reduce heat build-up and denaturation
* precaution: do on ice to reduce heat build-up and denaturation
material: drills (polytron), pestle/tube (dounce) bead beaters


=== Freeze-thaw ===
=== Freeze-thaw ===
* least effective method
* least effective method
* plus: does not denature proteins as much as other methods
* plus: does not denature proteins as much as other methods
material: pestle and mortar, liquid nitrogen


=== Detergents ===
=== Detergents ===
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* problems: detergent may inhibit subsequent reactions
* problems: detergent may inhibit subsequent reactions
* problems: detergent may disrupts protein interactions
* problems: detergent may disrupts protein interactions
no additional material required, just the chemicals and typical tubes


== Specific protocols ==
== Specific protocols ==
* [[Silver: Lysate for Western]] - Silver Lab: cell lysis for [[Western]] (method: Triton)
 
* [[Sauer:Lysing E. coli with Lysozymes]] - Sauer Lab: bacteria lysis (method: lysozyme)
{| {{sorttable}}
* [[Blackburn:Yeast Colony PCR]] - Blackburn Lab: yeast lysis for [[colony PCR]] (method NaOH)
! style="background:lightgrey"|description/link
* [[Jacobs:Protocol Total Protein Isolation Using RIPA Lysis Buffer]] - Jacobs Lab: cell line lysis for protein (method: RIPA buffer)
! style="background:lightgrey"|target
* [[Streptomyces:Protocols/Mini-Maxi Prep]] - Streptomyces community, East Anglia: bacteria lysis for DNA (method: NaOH)
! style="background:lightgrey"|lysis method
! style="background:lightgrey"|type of material
|-
| [[Silver: Lysate for Western]]
| protein
| detergent (Triton)
| cells
|-
| [[Sauer:Lysing E. coli with Lysozymes]]
|
| lysozyme
| bacteria
|-
| [[Blackburn:Yeast Colony PCR]]
| DNA
| NaOH
| yeast
|-
| [[Jacobs:Protocol Total Protein Isolation Using RIPA Lysis Buffer]]
| protein
| detergent (RIPA buffer)
| cells
|-
| [[Streptomyces:Protocols/Mini-Maxi Prep]]
| DNA
| NaOH
| bacteria
|-
| [[Western Blot/Tissue Preparation]]
| protein
| detergent & physical disruption
| tissue
|-
| [[Eccles:Protein Lysates from Cells in Culture]]
| protein
| detergent (SDS, NP40)
| cells
|-
| [[Eccles:Protein Lysates from Tissue]]
| protein
| detergent, physical, freezing
| tissue
|-
| [[RNA extraction using trizol/tri]]
| RNA
| phenol
| cells
|-
| [[Sauer:RNA Purification from E. coli]]
| RNA
| various options
| bacteria
|-
| add another method's name
| target
| lysis method
| type of starting material
|}
 
== Related hub pages ==
* [[RNA extraction]]


== Related discussions on BioForum ==
== Related discussions on BioForum ==
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[[Category:Protocol]][[Category:Material]]
[[Category:Protocol]][[Category:DNA]][[Category:RNA]][[Category:protein]]
[[Category:Escherichia]]
[[Category:Material]][[Category:Chemical]]
[[Category:Escherichia coli]][[Category:Mouse]][[Category:Streptomyces]][[Category:Yeast]]

Revision as of 03:45, 19 December 2008

This page should point you to the many different general and lab-specific protocols describing tissue and cell lysis and serve as forum for comparison. The ups and downs of various methods from sonication, homogenization, freeze-thaw cycles, and detergent-based lysis are examined below.

Comparison of lysis methods

Sonication

  • most efficient method of cell fractionation
  • problem: heat build up which can denature proteins (proportionate to length of sonication)
  • precaution: do on ice and sonicate intermittantly

material: ultrasonication bath or rods

Homogenisation

  • best for animal tissue; less suitable for cells
  • precaution: do on ice to reduce heat build-up and denaturation

material: drills (polytron), pestle/tube (dounce) bead beaters

Freeze-thaw

  • least effective method
  • plus: does not denature proteins as much as other methods

material: pestle and mortar, liquid nitrogen

Detergents

  • chemical method of lysis
  • problems: detergent may inhibit subsequent reactions
  • problems: detergent may disrupts protein interactions

no additional material required, just the chemicals and typical tubes

Specific protocols

description/link target lysis method type of material
Silver: Lysate for Western protein detergent (Triton) cells
Sauer:Lysing E. coli with Lysozymes lysozyme bacteria
Blackburn:Yeast Colony PCR DNA NaOH yeast
Jacobs:Protocol Total Protein Isolation Using RIPA Lysis Buffer protein detergent (RIPA buffer) cells
Streptomyces:Protocols/Mini-Maxi Prep DNA NaOH bacteria
Western Blot/Tissue Preparation protein detergent & physical disruption tissue
Eccles:Protein Lysates from Cells in Culture protein detergent (SDS, NP40) cells
Eccles:Protein Lysates from Tissue protein detergent, physical, freezing tissue
RNA extraction using trizol/tri RNA phenol cells
Sauer:RNA Purification from E. coli RNA various options bacteria
add another method's name target lysis method type of starting material

Related hub pages

Related discussions on BioForum

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