Collagenase

Source:
Clostridium histolyticum
CAS:
9001-12-1
EC:
3.4.24.3

Collagenase Raw Material Certification 

 

Crude collagenase preparations contain several isoforms of two different collagenases, a sulfhydryl protease, clostripain, a trypsin-like enzyme, and an aminopeptidase. This combination of collagenolytic and proteolytic activities is effective at breaking down intercellular matrices, the essential part of tissue dissociation. One component of the complex is a hydrolytic enzyme which degrades the helical regions in native collagen preferentially at the Y-Gly bond in the sequence Pro-Y-Gly-Pro, where Y is most frequently a neutral amino acid. This cleavage yields products susceptible to further peptidase digestion. Crude collagenase is inhibited by metal chelating agents such as cysteine, EDTA or o-phenanthroline but not DFP. It is also inhibited by α2-macroglobulin, a large plasma glycoprotein. Ca2+ is required for enzyme activity. Particular enzymatic profiles of each collagenase have been correlated with the tissues from which the cells for study were obtained (or with the uses to which the cells are put) and as a result of the correlations several types of crude collagenases have been established by Worthington: Types 1, 2, 3, and 4.

Type 1 crude collagenase has the original balance of collagenase, caseinase, clostripain and tryptic activities.
Type 2 contains higher relative levels of protease activity, particularly clostripain.
Type 3 contains lowest levels of secondary proteases.
Type 4 is designed to be especially low in tryptic activity to limit damage to membrane proteins and receptors.
• Type 5 contains higher collagenase and caseinase values.
• Type 6 contains high collagenase activity with a caseinase to collagenase ratio ~2:1. Designed to be enriched for Type II (col H) collagenase relative to Type I (col G).
• Type 7 contains collagenase and caseinase activities four-fold higher than collagenase Types 1 and 2.

 • Purified collagenase, Code: CLSPA, contains minimal secondary proteolytic activities along with high collagenase activity.

 

NEW! Animal Free Types AFA, AFB and AFC, STZ1, and STZ2 collagenases are derived from cultures grown in medium completely devoid of animal based components and designed for bioprocessing applications where introduction of potential animal derived pathogens must be prevented. Levels of secondary proteases are similar to Types 1 and 2 collagenase.

• CLSAFA is the original AFA grade designed to have collagenase and secondary proteases similar to Types 1 and 2 collagenase.
• CLSAFB contains higher collagenase and caseinase activities than CLSAFA.
• CLSAFC has especially low tryptic activity similar to Type 4 collagenase.
• STZ1 & STZ2, 0.22μ filtered STEMxyme® AOF Collagenase/Neutral Protease (Dispase®) blends for primary and stem cell isolation. 

Worthington also offers 0.22µm filtered preparations of each type in pre-packaged form for direct reconstitution and use in cell isolation and culture procedures. Correlations between enzyme type and effectiveness with different tissues have been good, but not perfect, due in part to the variable parameters of use. Nevertheless most researchers consider the tissue-typing of crude collagenase lots to be a valuable service. A detailed description of the Worthington collagenase and contaminant assays can be found in the Worthington Enzyme Manual. In addition, tissue specific references and detailed isolation conditions can be found in the Worthington Tissue Dissociation Guide.

The collagenase assay is a modification of Mandl wherein collagenase is incubated for five hours with native collagen and the extent of collagen breakdown is determined using the Moore and Stein, JBC, 176, 367, (1948) colorimetric ninhydrin method. Amino acids released are expressed as micromoles leucine per milligram collagenase.

Uses: Crude collagenases are widely used in enzymatic primary cell isolation and tissue dissociation procedures. Most researchers employ either crude collagenase preparations such as Types 1, 2, 3, and 4 or chromatographically purified collagenase (Code: CLSPA); the latter usually combined with secondary enzymes such as elastase, hyaluronidase, etc. For best results, the precise mixture of proteolytic activities must be tailored to the tissue to be dissociated. Correlations between type and effectiveness with different tissues have been good, but not perfect, and may be dependent partly on parameters of use and objectives as well as lot-to-lot variations. For more information see the Collagenase Sampling Program information. Worthington also publishes a Tissue Dissociation Guide which provides additional information regarding the enzymes used for these applications and specific references for numerous cell and tissue types.

Related Products:

Collagen (CL)
Deoxyribonuclease I (DP/D/DCLS/D2/DPFF/DPRF)
Elastase (ES/ESL/ESFF)
Hyaluronidase (HSE/HSEP)
Neutral Protease (Dispase, NPRO)
Papain (PAP/PAPL/PAP2)Proteinase K (PROK)
Trypsin (TL/TRL/TRL3/TRLS/TRTPCK)
Trypsin Inhibitors (LBI/OI/SI/SIC)

Cell Isolation Optimizing System (CIT)
Hepatocyte Isolation System (HIS)
Neonatal Cardiomyocyte Isolation System (NCIS)
Papain Dissociiation System (PDS/PDS2)

Unit Definition

One Unit releases one micromole of L-leucine equivalents from collagen in 5 hours at 37°C, pH 7.5

Collagenase Products

Description
Activity
Code
Cat. #
Price
Size
Description
Collagenase, Purified
Source:
Clostridium histolyticum
Chromatographically purified. ≤ 50 caseinase units per milligram. Supplied as a lyophilized powder.
Store at 2-8°C.
Activity
≥500 units per mg dry weight
Code
CLSPA
Cat. #
LS005275
Description
Collagenase Vial, NCIS
Source:
Clostridium histolyticum
A component of the NCIS kit. This material is 0.22 micron membrane filtered and lyophilized in autoclaved vials. A vial reconstituted with 5 ml of HBSS or equivalent yields a solution of 300 units/ml of collagenase, Code: CLSPA. Suitable for cell isolation and culture applications.
Store at 2-8°C.
Activity
≥500 units per mg dry weight
Code
CLSPANK
Cat. #
LK003240
Description
Collagenase/Elastase Vial (CLSH)
Source:
Clostridium histolyticum
Enzyme vial of HIS kit that contains 20,000 units of collagenase (Code: CLS-1) and 30 units of elastase (Code: ESL), filtered through 0.22 micron membrane and lyophilized.
Store at 2-8°C.
Activity
≥20,000 units collagense/30 units elastase per vial
Code
CLSH
Cat. #
LK002067
Description
Collagenase, Type 1
Source:
Clostridium histolyticum
The original balance of enzymatic activities. Each lot assayed for collagenase, caseinase, clostripain and tryptic activities. Suggested for epithelial, liver, lung and adrenal primary cell isolations. A dialyzed, lyophilized powder.
Store at 2-8°C.
Activity
≥125 units per mg dry weight
Code
CLS-1
Cat. #
LS004194
Description
Collagenase, Type 2
Source:
Clostridium histolyticum
Prepared to contain higher clostripain activity. Suggested for bone, heart, liver, thyroid and salivary primary cell isolation. Supplied as a dialyzed, lyophilized powder.
Store at 2-8°C.
Activity
≥125 units per mg dry weight
Code
CLS-2
Cat. #
LS004174
Description
Collagenase, Type 3
Source:
Clostridium histolyticum
Lower in secondary proteolytic contaminant activities but with typical collagenase activity. Suggested for mammary primary cell isolation. A dialyzed, lyophilized powder.
Store at 2-8°C.
Activity
≥100 units per mg dry weight
Code
CLS-3
Cat. #
LS004180
Description
Collagenase, Type 4
Source:
Clostridium histolyticum
Prepared to contain lower tryptic activity levels to limit damage to membrane proteins and receptors but with normal to above normal collagenase activity. Suggested for pancreatic islet primary isolation. A dialyzed, lyophilized powder.
Store at 2-8°C.
Activity
≥160 units per mg dry weight
Code
CLS-4
Cat. #
LS004186
Description
Collagenase, Type 5
Source:
Clostridium histolyticum
Partially purified to contain higher collagenase and caseinase activities. A dialyzed, lyophilized powder.
Store at 2-8°C.
Activity
≥450 units per mg dry weight
Code
CLS-5
Cat. #
LS005280
Description
Collagenase, Type 6
Source:
Clostridium histolyticum
Prepared to contain above normal collagenase, FALGPA and Caseinase activities and designed to be enriched with Type II (col H) collagenase.
Store at 2-8°C.
Activity
≥400 units per milligram dry weight
Code
CLS-6
Cat. #
LS005318
Description
Collagenase, Type 7
Source:
Clostridium histolyticum
Prepared to contain 4-5X more collagenase, FALGPA and Caseinase activities than Collagenase 1-4 and with moderate clostripain and tryptic activities.
 
 
Store at 2-8°C.
Activity
≥1,000 units per milligram dry weight
Code
CLS-7
Cat. #
LS005332
Description
Collagenase, Type 1, Filtered
Source:
Clostridium histolyticum
Collagenase, Type 1 (Code: CLS-1), which is filtered through a 0.22 micron membrane and lyophilized in vials.
Store at 2-8°C.
Activity
≥125 units per mg dry weight
Code
CLSS-1
Cat. #
LS004214
Description
Collagenase, Type 2, Filtered
Source:
Clostridium histolyticum
Collagenase, Type 2 (Code: CLS-2), which is filtered through a 0.22 micron membrane and lyophilized in vials.
Store at 2-8°C.
Activity
≥125 units per mg dry weight
Code
CLSS-2
Cat. #
LS004202
Description
Collagenase, Type 3, Filtered
Source:
Clostridium histolyticum
Collagenase, Type 3 (Code: CLS-3), which is filtered through a 0.22 micron membrane and lyophilized in vials to contain 50 milligrams per vial.
Store at 2-8°C.
Activity
≥100 units per mg dry weight
Code
CLSS-3
Cat. #
LS004206
Description
Collagenase, Type 4, Filtered
Source:
Clostridium histolyticum
Collagenase, Type 4 (Code: CLS-4), which is filtered through a 0.22 micron membrane and lyophilized in vials.
Store at 2-8°C.
Activity
≥160 units per mg dry weight
Code
CLSS-4
Cat. #
LS004210
Description
Collagenase, Type 5, Filtered
Source:
Clostridium histolyticum
Collagenase Type 5 (Code: CLS-5), which is filtered through a 0.22 micron membrane and lyophilized in vials.
Store at 2-8°C.
Activity
≥450 units per mg dry weight
Code
CLSS-5
Cat. #
LS005286
Description
Collagenase CLSAFA Animal Free/AF
Source:
Clostridium histolyticum
Prepared cultures grown in medium completely devoid of animal based components and designed for bioprocessing applications where introduction of animal derived pathogens must be prevented.
Store at 2-8°C.
Activity
≥150 units per mg dry weight
Code
CLSAFA
Cat. #
LS004152
Description
Collagenase, CLSAFA, Filtered
Source:
Clostridium histolyticum
Collagenase, Animal Free/AF (Code: CLSAFA) which is filtered through a 0.22 micron membrane and lyophilized in vials.
Store at 2-8°C.
Activity
≥150 units per mg dry weight
Code
CLSAFAS
Cat. #
LS004118
Description
Collagenase CLSAFB Animal Free/AF
Source:
Clostridium histolyticum
Prepared from cultures grown in medium completely devoid of animal based components and designed for bioprocessing applications where introduction of animal derived pathogens must be prevented.
Store at 2-8°C.
Activity
≥300 units per mg dry weight
Code
CLSAFB
Cat. #
LS004145
Description
Collagenase, CLSAFB, Filtered
Source:
Clostridium histolyticum
Collagenase, Animal Free/AF (Code: CLSAFB) which is filtered through a 0.22 micron membrane and lyophilized in vials.
Store at 2-8°C.
Activity
≥300 units per mg dry weight
Code
CLSAFBS
Cat. #
LS004124
Description
Collagenase CLSAFC Animal Free/AF
Source:
Clostridium histolyticum
Prepared from cultures grown in medium completely devoid of animal based components and designed for bioprocessing applications where introduction of animal derived pathogens must be prevented.
Store at 2-8°C.
Activity
≥200 units per mg dry weight
Code
CLSAFC
Cat. #
LS004138
Description
Collagenase, CLSAFC, Filtered
Source:
Clostridium histolyticum
Collagenase Animal Free/AF (Code: CLSAFC) which is filtered through a 0.22 micron membrane and lyophilized in vials.
Store at 2-8°C.
Activity
≥200 units per mg dry weight
Code
CLSAFCS
Cat. #
LS004130
Description
STEMxyme® 1, Collagenase/Neutral Protease (Dispase)
Source:
Clostridium histolyticum/Bacillus polymyxa
A specialized combination of Animal Free Clostridium histolyticum collagenase (Code: CLSAFB) and animal free Bacillus polymyxa neutral protease with a minimum of 250 CLS units and 1,000 caseinase units per mg dry weight. Designed for stem cell and other primary cell isolations and bioprocessing applications where introduction of potential animal derived pathogens must be prevented.
Store at 2-8°C.
Activity
≥250 CLS/1,000 CAS units per milligram dry weight
Code
STZ1
Cat. #
LS004106
Description
STEMxyme® 2 Collagenase/Neutral Protease (Dispase)
Source:
Clostridium histolyticum/Bacillus polymyxa
A specialized combination of Animal Free Clostridium histolyticum collagenase (Code: CLSAFB) and animal free Bacillus polymyxa neutral protease with a minimum of 250 CLS units and 2,000 caseinase units per mg dry weight. Designed for stem cell and other primary cell isolations and bioprocessing applications where introduction of potential animal derived pathogens must be prevented.
Store at 2-8°C.
Activity
≥250 CLS/2,000 CAS units per milligram dry weight
Code
STZ2
Cat. #
LS004112