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Deoxyribonuclease I - Assay

Source:
Bovine Pancreas
CAS:
9003-98-9
EC:
3.1.21.1
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Deoxyribonuclease I Assay

Method
That developed by Kunitz (1950) based upon the increased absorbance at 260 nm observed during the depolymerization of DNA by DNase. A unit causes an increase in absorbance at 260 nm of 0.001 per minute per ml when acting upon highly polymerized DNA at 25deg.C and pH 5.0 under the specified conditions. A standard enzyme preparation should be run in parallel with an unknown because standardization of DNA preparations and their degree of polymerization in solution is not possible.
Reagents
  • 1.0 M Acetate buffer, pH 5.0
  • 6.25 mM Magnesium sulfate in reagent grade water
  • Worthington Standard DNase Vial (Code:DSV) containing a defined activity of approximately 2000 DNase units per vial.
  • Worthington Highly Polymerized DNA (Code: DNA). Dissolve 10 mg DNA in 200 ml of 6.25 mM magnesium sulfate. Let stand overnight at room temperature. Add 25 ml of 1.0 M acetate buffer, pH 5.0 and dilute to a final volume of 250 ml with reagent grade water. (Substrate solution may be prepared in larger batches and stored for 2-3 weeks at 0 - 4deg.C.)
Enzyme

Note: Pancreatic deoxyribonuclease is unusually sensitive to physical denaturation by shaking. Mixing should be done by gentle inversion. Dissolve the standard vial in 1.0 ml of reagent grade water. Care must be taken when opening the vial that no lyophilized material is lost. This solution will contain the number of units/ml as stated on the label. Dilute further to a concentration of 20-60 u/ml. All dilutions are made in reagent grade water.

Sample to be assayed: Dissolve at a concentration of 1 mg/ml. Dilute further to a concentration of 20-60 u/ml immediately before the assay.

Procedure

Adjust spectrophotometer at 260 nm and 25deg.C. Pipette 2.5 ml of substrate into cuvettes and incubate in spectrophotometer at 25deg.C for 3-4 minutes to establish blank rate if any, and to reach temperature equilibration. Add 0.5 ml of diluted standard and record A260 for 8 - 10 minutes. Calculate ≥A260/minute from linear portion of curve following a brief lag.

Note: The change in A260 for this assay is not generally linear from the initial time and is linear for only short periods. The most linear portion should be used in determining the activity. A rate of 0.008 - 0.018 ≥[[Alpha]]/min. is recommended.

Calculate the "factor" for the standard vial.

equation

Using the diluted sample to be tested, repeat the above procedure. Record the ≥A260/minute from the linear portion of the curve.

Calculation

Activity is compared to that of the standard vial. Deoxyribonuclease activity can also be conveniently measured in a radial diffusion assay system.

Deoxyribonuclease I Products

Description
Activity
Code
Cat. #
Size
Price
Description
Deoxyribonuclease I, RNase & Protease Free, Solution
Source:
Bovine Pancreas
Molecular Biology Grade. Chromatographically purified to remove RNase and protease. Supplied as a solution at approximately 2 Kunitz units per microliter (approximately 1 mg/ml) containing 50% glycerol and 1mM calcium chloride.
Store at -20°C.
…
Activity
≥2,000 Kunitz units per ml
Code
DPRFS
LS006342
100 un
$32.00
LS006344
500 un
$90.00
LS006348
Bulk
---
The minimum amount for bulk packaging/pricing for this product is 20000 x 100 un. Please contact custservice@worthington-biochem.com to request a quote for smaller amounts.
Description
Deoxyribonuclease I, Ribonuclease&Protease Free
Source:
Bovine Pancreas
Molecular Biology Grade. Chromatographically purified to remove RNase and protease. Lyophilized in vial containing glycine and calcium as stabilizer.
Store at 2-8°C. PROTECT FROM MOISTURE.
…
Activity
≥2,000 Kunitz units per mg dry weight
Code
DPRF
LS006331
2500 un
$50.00
LS006333
10 ku
$176.00
LS006343
50 ku
$640.00
LS006334
Bulk
---
The minimum amount for bulk packaging/pricing for this product is 2000000 x 1 un. Please contact custservice@worthington-biochem.com to request a quote for smaller amounts.
Description
Deoxyribonuclease I
Source:
Bovine Pancreas
Chromatographically purified. A lyophilized powder containing glycine as a stabilizer. Contains ≤0.0005% RNase.
Store at 2-8°C. PROTECT FROM MOISTURE.
…
Activity
≥2,000 Kunitz units per mg dry weight
Code
DPFF
LS006330
25 ku
$102.00
LS006328
125 ku
$375.00
LS006332
Bulk
---
The minimum amount for bulk packaging/pricing for this product is 2500 x 1 ku. Please contact custservice@worthington-biochem.com to request a quote for smaller amounts.
Description
Deoxyribonuclease I
Source:
Bovine Pancreas
Chromatographically purified. A lyophilized powder with glycine as a stabilizer.
Store at 2-8°C. PROTECT FROM MOISTURE.
…
Activity
≥2,000 Kunitz units per mg dry weight
Code
D
LS002004
5 mg
$40.00
LS002006
20 mg
$96.00
LS002007
100 mg
$350.00
LS002009
Bulk
---
The minimum amount for bulk packaging/pricing for this product is 2000 x 1 mg. Please contact custservice@worthington-biochem.com to request a quote for smaller amounts.
Description
Deoxyribonuclease I, Filtered
Source:
Bovine Pancreas
Filtered through a 0.22 micron membrane and lyophilized in vials. Material is not tested for pyrogenicity.
Store at 2-8°C. PROTECT FROM MOISTURE.
…
Activity
≥2,000 Kunitz units per mg dry weight
Code
DCLS
LS002058
11 mg
$110.00
LS002060
25 mg
$210.00
Description
Deoxyribonuclease I, Standard Vial
Source:
Bovine Pancreas
Lyophilized in vials for assay standardization. Labeled to show established activity. Not suitable for assays at neutral pH.
Store at 2-8°C.
…
Activity
~2,000 Kunitz units per vial
Code
DSV
LS002173
2 ku
$23.00
LS002172
5x2 ku
$66.00
Description
DNase Vial (D2)
Source:
Bovine Pancreas
A component of the Papain Dissociation System. This material is 0.22 micron membrane filtered and lyophilized in autoclaved vials. A vial reconstituted with 0.5 ml of EBSS or equivalent yields a solution of 2000 units/ml of deoxyribonuclease (1 mg/ml).
Store at 2-8°C.
…
Activity
≥1,000 Kunitz units per vial
Code
D2
LK003170
1 vi
$28.00
LK003172
5 vi
$92.00
Description
Deoxyribonuclease I
Source:
Bovine Pancreas
Partially purified. A lyophilized powder.
Store at 2-8°C.
…
Activity
≥2,000 Kunitz units per mg dry weight
Code
DP
LS002138
25 mg
$45.00
LS002139
100 mg
$110.00
LS002140
1 gm
$900.00
LS002141
Bulk
---
The minimum amount for bulk packaging/pricing for this product is 20000 x 1 mg. Please contact custservice@worthington-biochem.com to request a quote for smaller amounts.
Description
Deoxyribonuclease I
Source:
Bovine Pancreas
Partially purified. Supplied as lyophilized powder.
Store at 2-8°C.
…
Activity
≥1,250 Kunitz units per mg dry weight
Code
DPB
LS002145
100 mg
$92.00
LS002147
1 gm
$675.00
LS002149
Bulk
---
The minimum amount for bulk packaging/pricing for this product is 20000 x 1 mg. Please contact custservice@worthington-biochem.com to request a quote for smaller amounts.
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