Papain Assay

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Papain Assay

Method

A titrimetric determinatin of the acid produced during the hydrolysis of benzoyl-L-arginine ethyl ester (BAEE). One unit will hydrolyze one micromole of benzoyl-L-arginine ethyl ester per minute at 25°C and pH 6.2 under the specified conditions.

Reagents

Enzyme diluent (Activation buffer): Prepare fresh daily by mixing the following:

0.01 M EDTA 10 ml
0.06 M Mercaptoethanol 0.1 ml
0.05 M Cysteine⋅HCl 10 ml
Reagent grade water 70 ml
Substrate solution: Prepare fresh daily by mixing the following:

0.058 M BAEE 15.0 ml
0.01 M EDTA 0.8 ml
0.05 M Cysteine⋅HCl 0.8 ml
Adjust pH to 6.2 and dilute to a final volume of 21 ml with reagent grade water.
Titrant: 0.01-0.02 N NaOH, standardized

Enzyme

Activate enzyme by dissolving in enzyme diluent to a concentration of 0.05-0.1 mg/ml. Under these conditions activation is complete within 30 minutes.

Determination of protein concentration

Procedure

The reaction can be measured with either an automatic titrator or a laboratory pH meter. The titration vessel should be maintained at 25°C.

Pipette the following into the titration vessel at 25°C:

Substrate solution 5.0 ml
3.0 M NaCl 5.0 ml
Reagent grade water 5.0 ml

At zero time add 0.1 ml of appropriately diluted enzyme and adjust the pH to 6.2. Record the amount of standardized NaOH added per minute to maintain the pH at 6.2 after a constant rate is achieved.

Calculation

\frac{\text{Units}}{\text{mg}}\text{ = } \frac{\frac{\text{ml base added}}{\text{min}}\text{ x normality x 1000}}{\text{mg enzyme in reaction mixture}}

Papain Products

Description

Papain, Suspension

Source:

Carica papaya Latex

Supplied as a two times crystalline suspension in 50mM sodium acetate, pH 4.5. To ensure full activity, the enzyme should be incubated in a solution containing 1.1mM EDTA, 0.067mM mercaptoethanol and 5.5mM cysteine-HCl for 30 minutes. It is recommended that the enzyme be 0.22 micron filtered after dissolution and prior to use.

Store at 2-8°C.

…

Ice Pack required

Activity

Activates to at least 20 units per mg protein

Certificate of Analysis

Code

PAP

Product details

LS003124

25 mg

$40.00

LS003126

100 mg

$88.00

LS003127

1 gm

$595.00

LS003128

Bulk

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Description

Papain, Lyophilized

Source:

Carica papaya Latex

Supplied as a lyophilized powder prepared from a two times crystalline suspension, Code: PAP. To ensure full activity, the enzyme should be incubated in a solution containing 1.1mM EDTA, 0.067mM mercaptoethanol and 5.5mM cysteine-HCl for 30 minutes. It is recommended that the enzyme be 0.22 micron filtered after dissolution and prior to use.

Store at 2-8°C.

…

Activity

Activates to at least 15 units per mg protein

Certificate of Analysis

Code

PAPL

Product details

LS003118

25 mg

$44.00

LS003119

100 mg

$100.00

LS003120

1 gm

$685.00

LS003122

Bulk

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Description

PDS Kit, Papain Vial

Source:

Papaya Latex

A component of the Papain Dissociation System, for use in the tissue dissociation method of Huettner, J.E., and Baughman, R.W.: J. Neuroscience, 6, 3044 (1986). Contains papain, L-cysteine, and EDTA. This material is 0.22 micron membrane filtered and lyophilized in autoclaved vials. A vial reconstituted with 5 ml of EBSS or equivalent yields a solution at 20 units of papain per ml in 1mM L-cysteine with 0.5mM EDTA.

Store at 2-8°C.

…

Activity

≥100 units per vial

Certificate of Analysis

Code

PAP2

Product details

LK003176

1 vi

$29.00

LK003178

5 vi

$98.00

0.01 M EDTA	10 ml
0.06 M Mercaptoethanol	0.1 ml
0.05 M Cysteine⋅HCl	10 ml
Reagent grade water	70 ml

0.058 M BAEE	15.0 ml
0.01 M EDTA	0.8 ml
0.05 M Cysteine⋅HCl	0.8 ml

Substrate solution	5.0 ml
3.0 M NaCl	5.0 ml
Reagent grade water	5.0 ml

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Papain - Assay

Papain Assay

Papain Products