Other Cells

General Approach

Recommended products: Defined Enriched Collagenases

Use our Defined Enriched (DE) Collagenase products as replacements for the crude, enriched, or purified collagenase products listed in the table below.

Table 1: Recommended Products

Product Name & Description

VitaCyte Equivalent

For every mg of collagenase product use following mg of DE Collagenase ± BP Protease products

Worthington
Type 1: contains average amounts of assayed activities DE 10 or 100  1
Type 2: contains greater clostripain activity than Type 1 DE 10 or 100 1
Type 3: contains lower protease activity than type 1 DE 20 or 200 0.65
Type 4 contains lower protease and tryptic activity (clostripain) than Type 1 DE 20 or 200 0.65
Type V contains higher collagenase and protease activity DE 10 or 100

DE 20 or 200

1
Sigma
Type I: similar to Worthington Type I but eith higher collagenase activity DE 10 or 100  1
Type IA: similar to Worthington Type I but eith higher collagenase activity DE 10 or 100 1
Type II: similar to Worthington Type II but eith higher collagenase activity DE 20 or 200 0.65
Type III: purified collagenase with trace amounts of protease Collagenase Gold 1
Type IV: lot tested for release of rat hepatocytes DE 10 or 100

DE 60 or 600

 

Follow protocol for rodent hepatocytes
Type V: lot tested for release of rat islets DE 40 or 400

DE 60 or 600

Follow protocol for rodent islets
Type VII: purified collagenase substantially free of protease and clostripain Collagenase Gold Contact VitaCyte for recommendation
Type VIII: tested for release of rat epididymal adipocytes or hepatoctyes DE 10 or 100

DE 60 or 600

1

Follow protocol for rodent hepatocytes

Type XI enriched collagenase prepared for islet isolation DE 60 or 600 1
 Manufactured by Roche; distributed by Sigma  
Collagenase P DE 20 or 200  1
 Liberase TL Research Grade  DE 40 or 400 1
 Liberase TM Research Grade  DE 10 or 100 2
Liberase TH Research Grade DE 100  Contact VitaCyte for recommendation
 Liberase DL Research Grade DE 80 or 800 1
Liberase DH Research Grade DE 20 or 200 1

Note: Sigma Type IV, V, and VII are function tested lots of crude collagenase for isolation of the stated cell types. In these cases, you are referred to use of the protocols for these cells that can be found in Applied Use.

Recommended method: Varies

Recommended method is dependent on the user’s experience.

Expected cell yield

Expected results are dependent on the user’s experience.

Perspectives

Crude collagenase has been used for many years to isolate a wide variety of cells. The major disadvantage for using recommendations provided by other manufacturer’s or results published in the scientific literature is that cell function or yields are dependent on the unique characteristics of the lot of crude collagenase used in the isolation procedure. The results obtained by one investigator may not be the same as those obtained by another because of the lot to lot variability of crude collagenase. All crude collagenase products are minimally purified, C. histolyticum culture supernatants whose composition reflects the biochemical composition at the end of the fermentation process.

VitaCyte’s low cost, DE Collagenase is designed to overcome many of the limitations of traditional collagenase products (i.e., crude or enriched collagenase) as shown in the table below.

Table 2: Comparison of Traditional Collagenase and VitaCyte DE Collagenase Products

Traditional Collagenase

DE Collagenase

Enzyme composition

Unknown

  • Minimally purified bacterial culture supernatant
  • Each lot has multiple proteases, each with different activities
  • Each lot has variable collagen degradation activity

 

Known

  • Increasing amounts of enriched collagenase (> 85% pure) with minimal protease activity add to fixed amount of purified BP Protease
  • Enriched collagenase has high collagen degradation activity and very low contamination with protease (<0.5%)
Need for lot pre-qualification

Strongly recommended

  • Dependent on user’s need to tightly control the reproducibility of the cell isolation process

 

 

 

Only on first use of product

  • Must determine optimal DE Collagenase product and dose on initial use
  • Once lot qualified, use same product with enzyme activities per mL in enzyme solution as guide to adjust mg of product per mL with next lot of product
Degree of experimental control

Low

  • Only variable under control is dose of enzyme used in the isolation procedure
  • Since each lot unique with unknown biochemical composition, it may be difficult to replicate results with new lot of product

High

  • Can control enzyme composition and dose used in isolation procedure
  • Can explore other enzyme formulations by using enriched collagenase (Collagenase Gold) in combination with BP Protease or other proteases
Translation of protocol to clinical use

Difficult

  • See comments on Degree of Experimental Control, difficult to prepare clinical grade product if do not know enzyme composition

Straightforward

  • Enzyme composition of DE products known; VitaCyte can translate these formulation to manufacture a clinical grade product

Resources & Reviews

For more information on any of these issues, request VitaCyte’s white paper on “Lot Qualification: A common but costly pathway to select enzymes for cell isolation” or a 33 min video of a tech talk by Bob McCarthy presented at the 2016 American Society of Cell Biology meeting: “Collagenase: Moving from black box to transparent knowledge”. The video reviews basic aspects of collagenase as it applies to cell isolation and presents an overview of VitaCyte’s DE Collagenase product line.