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Bioprocessing with Salt Active Nucleases  - High Salt Conditions

Bioprocessing with Salt Active Nucleases  - High Salt Conditions

Bioprocessing with Salt Active Nucleases  - High Salt Conditions

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For SAN HQ, SAN HQ ELISA Kit, and now SAN HQ GMP


SAN HQ GMP is biochemically identical to SAN HQ but produced under GMP conditions.

Jump to SAN HQ GMP section 👇

Applications

  • Purification of biologics from residual nucleic acids in biopharma manufacturing
  • Purification of recombinant proteins and enzymes for research and diagnostic use
  • Removal of unwanted nucleic acids contamination in molecular biology reagents in challenging conditions
  • Reduction of viscosity in biological samples during production and automation
  • Vaccine manufacturing and viral vector preparation
  • DNA removal in high-salt lysates

SAN HQ  - Peak performance at high salt conditions

Salt Active Nuclease High Quality (SAN HQ) is a Bioprocessing Grade nuclease developed as the most efficient solution for removal of both single and double stranded DNA and RNA at high salt conditions. 

This nonspecific endonuclease has peak activity at salt concentrations between 400 - 700 mM (Fig. 1)

Non-enveloped viruses like Adenoviruses and Adeno-Associated Viruses (AAV’s) are inherently more robust with two distinct advantages: 1) They exhibit higher tolerance to additives like salt and detergents and 2) their production often involves the lysis of host cells, allowing for harvesting non-secreted vectors. 

For Adeno-Associated Viruses (AAVs), which are often harvested from crude cell lysate, the high salt tolerance of SAN HQ is particularly beneficial. Salt is typically added to such lysates to reduce viral aggregation, facilitating more effective nuclease action to digest residual DNA.

SAN HQ's is engineered for optimum activity in these high salt environments ensuring that you achieve unparalleled DNA removal without compromising the integrity of these robust viral vectors.

Key Benefits

  • Optimized Residual DNA Removal: Ensures efficient degradation of residual DNA in high-salt conditions, meeting stringent quality requirements for biologics and vaccines.
  • Boosted AAV Vector Purification: Enhances the purification process for adeno-associated viral vectors in high-salt conditions, improving quality and yield.
  • Streamlined Workflow: Eliminates the need for desalting stages, simplifying the bioprocessing protocol and saving time and resources.
  • Enable High-Throughput Processes: Facilitates scale-up and automation by working effectively in high-salt environments, increasing operational throughput.
  • Potential Surge in Virus Yield: Operates under conditions that may boost the titer yield of AAV production, potentially enhancing overall viral yield.
  • Economized Enzyme Usage: Reduces the need for excess enzyme and additional process adjustments, resulting in significant cost savings.
  • Minimized Risk of Process Disruptions: Offers reliable performance in various high-salt bioprocessing conditions, reducing the likelihood of disruptions due to enzyme inhibition.
  • Reliability: Provides consistent enzyme activity in challenging high-salt conditions, adding a layer of predictability and dependability to your operations.
  • Broader Applicability: Versatile enough to be used in a wide range of viral vector systems, expanding your research and production capabilities.
  • Enhanced Viral Stability: High-salt levels stabilize viral vectors, and SAN HQ operates effectively in these conditions, maintaining high yield and quality.
  • Host Cell Lysis: Facilitates efficient lysis of host cells in high-salt conditions, optimizing the harvest of both secreted and non-secreted viral vectors.

Key Features 

  • High purity (≥ 98%)
  • No protease detected
  • Supplied with extended product documentation
  • Compatible with SAN HQ ELISA

The Challenge in Removing Host Cell Chromatin Impurities 

In bioprocessing, the primary role of a nuclease is to efficiently digest and fragment host-cell DNA into sufficiently small pieces, facilitating its removal during downstream processing. While most nucleases can effectively degrade naked DNA into tiny fragments under optimal conditions—as demonstrated by M-SAN HQ and SAN HQ, which can digest dsDNA into fragments smaller than 6 nt—the reality in bioprocessing is more complex. (See fig. 5)

The DNA targeted for removal often exists as chromatin, embedded in a complex matrix containing remnants of the lysed host cell as well as large amounts of the therapeutic product.The product may or may not have an affinity for the chromatin you aim to remove.

High salt is often applied to mitigate issues like aggregation. The real challenge lies in a nuclease's ability to efficiently fragment chromatin under these more complicated, high-salt, conditions—not merely degrading naked DNA under ideal circumstances.

See Case Study on Efficient Fragmentation: Superior Chromatin Digestion Before Flow-Through Purification

SAN HQ ELISA Kit

SAN HQ ELISA kit is developed for the detection and quantification of SAN HQ and SAN  HQ GMP. The kit is designed as a classical sandwich ELISA, with two monoclonal antibodies specific towards SAN HQ nuclease (fig 6).

Features

  • Sensitive: 0.4 – 25.6 ng/ml
  • Precise: RSD ≤ 15%
  • Accurate: 100% ± 15%
  • Stability: 12 months when stored between +2°C to +8°C

Figures

SAN HQ (High salt) - Optimum activity in solutions with high salinity
Figure 1. Optimum activity in solutions with high salinity
SAN HQ (High salt) - Optimum activity in solutions with high salinity

SAN High Quality has optimum activity between 400 - 700 mM NaCl but can be used in a broad range of NaCl-concentrations. The activity was measured at 37°C in a 25 mM Tris-HCl buffer, pH 8.5, 5 mM MgCl2 with varying concentrations of NaCl. Maximum activity was set to 100%. Similar results were obtained using KCl instead of NaCl (results not shown).

SAN HQ (High salt) - Optimum performance at slightly alkaline conditions
Figure 2. Optimum performance at slightly alkaline conditions
SAN HQ (High salt) - Optimum performance at slightly alkaline conditions

.

SAN HQ performs best in the 8.0 - 8.8 pH range. A working range starting at pH 7.0 allows use in most biological buffers. The activity was measured at 37°C in a 25 mM Tris-HCl or Bis-Tris Propane buffer buffer at various pH, 5 mM MgCl2 and 500 mM NaCl. Maximum activity was set to 100%.

SAN HQ (High salt) -  Suitable for DNA removal in the ambient to 37°C range
Figure 3. Suitable for DNA removal in the ambient to 37°C range
SAN HQ (High salt) - Suitable for DNA removal in the ambient to 37°C range

SAN HQ originates from a cold-adapted organism, which allows excellent performance at the temperature ranges commonly used in bioprocessing. For temperature sensitive products, DNA digestion overnight at 4°C is also possible. The activity was measured in a 25 mM Tris-HCl buffer at pH 8.5 (@ measurement temperature), 5 mM MgCl2 and 500 mM NaCl. 37°C was set to 100%.

SAN HQ (High salt) - Robust performance at 37°C
Figure 4. Robust performance at 37°C
SAN HQ (High salt) - Robust performance at 37°C

SAN HQ shows excellent stability during use at 37°C, which is the typical incubation temperature in bioprocessing. After incubation at relevant DNA digestion conditions for 1 hour at 37°C, no significant loss of activity was observed compared to control sample kept on ice in enzyme dilution buffer. Buffer compositions: Lysis buffer (50 mM Tris-HCl pH 8.0 @ 25°C, 5 mM MgCl2, 0.5 % Triton X-100, 500 mM NaCl), DMEM (DMEM supplemented with 10 % FBS, 5 mM MgCl2 and 300 mM NaCl), Lysis buffer in DMEM (Lysis buffer and DMEM 1:1).

SAN HQ (High salt) - Efficient Residual DNA Fragmentation
Figure 5. Efficient Residual DNA Fragmentation
SAN HQ (High salt) - Efficient Residual DNA Fragmentation

Gel analysis showing that SAN HQ and M-SAN HQ effectively reduces residual host cell DNA to under 6 nucleotides.

SAN HQ (High salt) - SAN HQ ELISA kit is ideal to quantify residual SAN HQ
Figure 6. SAN HQ ELISA kit is ideal to quantify residual SAN HQ
SAN HQ (High salt) - SAN HQ ELISA kit is ideal to quantify residual SAN HQ

The SAN HQ ELISA kit is designed as a classical sandwich ELISA. Two SAN HQ-specific antibodies recognising different epitopes are used to respectively capture and detect SAN HQ in test samples. The detection antibody is linked to Biotin, which is detected by Streptavidin in the next step. Streptavidin is linked  to a Peroxidase which catalyses a reaction that can be quantified colorimetrically  by a spectrophotometer measuring OD at 450 nm.

Properties

Specifications
Source

Recombinantly produced in Pichia pastoris

Molecular weight

24.8 kDa.
Molecular Weight is theoretically calculated from the peptide sequence

Protein Purity

≥ 98% by SDS-PAGE analysis

Isoelectric point

9.55

Unit Definition

One unit is defined as the amount of enzyme that causes a ΔA260 = 1.0 in 30 minutes at 37°C in 25 mM Tris-HCI pH 8.5 (@25°C), 5 mM MgCl2' 500 mM NaCl, and 50 μg/ml calf thymus DNA.

Specificity

Nonspecific endonuclease cleaving single and double stranded DNA and RNA.

Working ranges
  • Temperature: 5 – 38°C, 4°C overnight, optimal: 32 - 38°C
  • Salt concentration (NaCl / KCl): 0 – 1000 mM, optimal: 400 – 700 mM
  • Mg2+: >1 mM is required for activity, optimal: 6 - 50 mM
  • pH: 7.0 - 9.5, optimal: 8.0 - 8.8

Note: The working range is defined as above 10% activity and optimal range as above 80% activity.

Tolerance to typical buffer additives
  • Imidazole: 20% activity at 350 mM Imidazole
  • Glycerol: 20% activity at 35% Glycerol
  • Triton X-100: No reduction in activity (tested up to 15%)
  • Tween 20: No reduction in activity (tested up to 10%)
  • SDS: Not recommended
  • Urea: Not recommended
  • Reducing agents (e.g. DTT, TCEP): will result in inactivation
Quality Control

SAN HQ GMP

SAN HQ GMP is biochemically identical to SAN HQ but produced under GMP conditions.

Biologics intended for therapeutic use must adhere to strict requirements for ancillary and raw materials used in their GMP-compliant manufacture. The Salt Active Nuclease from ArcticZymes is intended for removal of unwanted DNA and RNA from any biological component manufactured using host cells.

With the introduction of GMP-grade Salt Active Nuclease, qualifying the use of this unique nuclease in GMP compliant manufacturing processes has been greatly simplified.

  • ISO 13485:2016 Certified Quality Management System
  • Manufactured under GMP conditions
  • Manufactured from Animal Origin Free (AOF) raw materials 
  • Regulatory support by U.S. FDA Drug Master File
  • Accompanied by ELISA kit for verifying removal of nuclease in DSP

Request a copy of our quality dossier for SAN HQ GMP  Request quality dossier here

To reference SAN HQ GMP Master file in your U.S. FDA application, please complete the request for a Letter of Authorization (LOA) Request LoA here

Recommended Protocols

Ordering Info

Quantity
Product Name
SKU
Pack Size
Concentration
Price
1
T7 RNA Polymerase
74100-201
10 000 U
50 U/µl
$ 145.00 USD
133.00
145.00
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2
T7 RNA Polymerase
74100-110
100 kU
≥ 500 U/µl
$ 0.00 USD
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3
T7 RNA Polymerase
74100-100
As per request
As per request
$ 0.00 USD
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4
T7 RNA Polymerase Glycerol Free
74110-110
100 kU
≥ 500 U/µl
$ 0.00 USD
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5
T7 RNA Polymerase Glycerol Free
74110-100
As per request
As per request
$ 0.00 USD
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1
T4 DNA Ligase
71800-202
250 kU
5000 U/μl
$ 32.00 USD
30.00
32.00
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1
AZscript RT
72100-201
10000 U
200 U/μl
$ 136.00 USD
123.00
136.00
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1
Heat&Run gDNA Removal Kit
80200-250
250 rxn
$ 651.00 USD
548.00
651.00
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1
M-SAN HQ ELISA Kit
70960-001
12x8 Strip plate
$ 756.00 USD
655.00
756.00
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1
HL-dsDNase Triton Free
70801-201
250 U
2 U/µl
$ 357.00 USD
305.00
357.00
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1
PCR Decontamination Kit
80400-100
100 rxn
$ 130.00 USD
110.00
130.00
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1
AZtaq™ DNA Polymerase
73100-201
500 U
5 U/µl
$ 83.00 USD
75.00
83.00
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1
HL-SAN Triton Free
70911-202
25000 U
25 U/µl
$ 560.00 USD
473.00
560.00
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1
ArcticZymes Proteinase
71600-201
50 U
>200 U/ml
$ 155.00 USD
131.00
155.00
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1
IsoPol®
71500-201
200 U
5 U/µl
$ 75.00 USD
68.00
75.00
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1
ArcticZymes R2D Ligase
71900-105
5 kU
10 U/µl
$ 419.00 USD
361.00
419.00
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1
rSAP
70700-201
1000 U
1 U/µl
$ 131.00 USD
116.00
131.00
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1
dsDNase Triton Free
70601-201
250 U
2 U/µl
$ 357.00 USD
305.00
357.00
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1
Cod UNG Triton Free
70501-201
100 U
1 U/µl
$ 130.00 USD
109.00
130.00
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1
IsoPol™ BST⁺
71502-202
1600 U
8 U/µl
$ 78.00 USD
71.00
78.00
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1
SAN High Quality
70920-150
500 kU
>250 U/µl
$ 5,767.00 USD
5011.00
5767.00
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1
HL-ExoI
70100-201
2500 U
>20 U/µl
$ 69.00 USD
65.00
69.00
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1
SAN HQ 2.0
70940-150
500 kU
250 U/µl
$ 5,767.00 USD
5011.00
5767.00
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2
Cod UNG Triton Free
70501-202
1000 U
1 U/µl
$ 548.00 USD
456.00
548.00
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2
SAN High Quality
70920-160
5 MU
>250 U/µl
$ 21,913.00 USD
19035.00
21913.00
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2
PCR Decontamination Kit
80400-500
500 rxn
$ 519.00 USD
445.00
519.00
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2
HL-dsDNase Triton Free
70801-202
1000 U
2 U/µl
$ 1,055.00 USD
877.00
1055.00
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2
IsoPol™ BST⁺
71502-120
10000 U
120 U/µl
$ 384.00 USD
350.00
384.00
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2
dsDNase Triton Free
70601-202
1000 U
2 U/µl
$ 1,055.00 USD
877.00
1055.00
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2
T4 DNA Ligase
71800-100
as per request
as per request
$ 0.00 USD
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2
ArcticZymes R2D Ligase
71900-100
As per request
As per request
$ 0.00 USD
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2
M-SAN HQ
70950-120
200 kU
>250 U/µl
$ 3,155.00 USD
3024.00
3155.00
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2
IsoPol® SD+
71501-201
200 U
5 U/µl
$ 75.00 USD
68.00
75.00
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2
HL-SAN Triton Free
70911-150
500 kU
25 U/µl
$ 5,425.00 USD
4714.00
5425.00
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2
rSAP
70700-202
5000 U
1 U/µl
$ 519.00 USD
450.00
519.00
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2
SAN HQ 2.0
70940-160
5 MU
>250 U/µl
$ 21,913.00 USD
19035.00
21913.00
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2
Heat&Run gDNA Removal Kit
80200-50
50 rxn
$ 166.00 USD
139.00
166.00
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2
HL-ExoI
70100-202
500 kU
>20 U/µl
$ 6,148.00 USD
5040.00
6148.00
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2
AZscript RT
72100-100
As per request
As per request
$ 0.00 USD
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2
ArcticZymes Proteinase
71600-110
1000 U
>200 U/ml
$ 1,419.00 USD
1188.00
1419.00
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2
AZtaq™ DNA Polymerase
73100-110
5000 U
>50 U/µl
$ 619.00 USD
568.00
619.00
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3
Cod UNG Triton Free
70501-110
10 kU
1 U/µl
$ 3,920.00 USD
3472.00
3920.00
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3
SAN HQ 2.0
70940-202
25 kU
25 - 30 U/µl
$ 577.00 USD
498.00
577.00
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3
rSAP
70700-203
25 kU
1 U/µl
$ 1,965.00 USD
1703.00
1965.00
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3
HL-ExoI
70100-150
50 kU
>20 U/µl
$ 683.00 USD
593.00
683.00
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3
M-SAN HQ
70950-150
500 kU
>250 U/µl
$ 6,055.00 USD
5803.00
6055.00
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3
ArcticZymes Proteinase
71600-100
As per request
>200 U/ml
$ 0.00 USD
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3
IsoPol® BST⁺ Glycerol Free
71512-100
As per request
As per request
$ 0.00 USD
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3
HL-dsDNase Triton Free
70801-203
2500 U
5 U/µ
$ 2,365.00 USD
1961.00
2365.00
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3
AZtaq™ DNA Polymerase
73100-100
As per request
As per request
$ 0.00 USD
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3
dsDNase Triton Free
70601-203
2500 U
5 U/µl
$ 2,365.00 USD
1961.00
2365.00
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3
SAN High Quality
70920-202
25 kU
25-30 U/µl
$ 577.00 USD
498.00
577.00
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3
HL-SAN
70910-202
25000 U
25 U/µl
$ 560.00 USD
473.00
560.00
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4
IsoPol® BST⁺ Glycerol HC Free
71522-100
As per request
As per request
$ 0.00 USD
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4
Cod UNG Triton Free
70501-151
50 kU
1 U/µl
$ 14,128.00 USD
12281.00
14128.00
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4
ArcticZymes Proteinase Glycerol-FREE
71610-100
As per request
>200 U/ml
$ 0.00 USD
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4
rSAP
70700-101
200 kU
1 U/µl
$ 13,375.00 USD
11631.00
13375.00
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4
dsDNase Triton Free
70601-150
50 kU
50 U/µl
$ 17,069.00 USD
14830.00
17069.00
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4
SAN HQ 2.0 ELISA Kit
70970-001
1x96 Well Plate
$ 755.00 USD
655.00
755.00
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4
HL-dsDNase Triton Free
70801-110
100 kU
2 U/µl
$ 45,552.00 USD
39608.00
45552.00
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4
HL-SAN
70910-150
500 kU
26 U/µl
$ 5,425.00 USD
4714.00
5425.00
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4
SAN High Quality GMP
70980-150
500 kU
400-600 U/ul
$ 0.00 USD
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5
Cod UNG Triton Free
70501-150
50 kU
50 U/µl
$ 14,128.00 USD
12281.00
14128.00
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5
HL-dsDNase Glycerol-Free - Triton Free
70811-50
5000 U
>50 U/µl
$ 4,031.00 USD
3505.00
4031.00
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5
rSAP Glycerol-free
70710-201
5000 U
>20 U/µl
$ 603.00 USD
521.00
603.00
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4
M-SAN HQ
70950-155
1 MU
>250 U/µl
$ 10,905.00 USD
10450.00
10905.00
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5
M-SAN HQ
70950-160
5 MU
>250 U/µl
$ 23,008.00 USD
22049.00
23008.00
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5
dsDNase Triton Free
70601-110
100 kU
2 U/µl
$ 25,604.00 USD
22260.00
25604.00
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5
SAN High Quality GMP
70980-160
5 MU
400-600 U/µl
$ 0.00 USD
Request Quote
6
rSAP Glycerol-free
70710-100
As per request
As per request
$ 0.00 USD
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6
Cod UNG Glycerol-Free Triton Free
70511-105
5000 U
50 U/µl
$ 2,598.00 USD
2256.00
2598.00
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6
dsDNase Glycerol Free, Triton Free
70611-50
5000 U
>50 U/µl
$ 4,031.00 USD
3505.00
4031.00
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6
SAN High Quality Triton Free
70921-150
500 kU
>250 U/µl
$ 5,767.00 USD
5011.00
5767.00
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6
M-SAN HQ
70950-202
25 kU
25-30 U/µl
$ 606.00 USD
580.00
606.00
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6
HL-dsDNase
70800-201
250 U
2 U/µl
$ 357.00 USD
305.00
357.00
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7
Cod UNG Glycerol-Free Triton Free
70511-150
50 kU
50 U/µl
$ 14,128.00 USD
12281.00
14128.00
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7
SAN High Quality Triton Free
70921-160
5 MU
>250 U/µl
$ 21,913.00 USD
19035.00
21913.00
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7
dsDNase
70600-201
250
2 U/µl
$ 357.00 USD
305.00
357.00
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7
HL-dsDNase
70800-202
1000 U
2 U/µl
$ 1,055.00 USD
877.00
1055.00
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8
SAN High Quality Triton Free
70921-202
25 kU
25-30 U/µl
$ 577.00 USD
498.00
577.00
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8
Cod UNG
70500-201
100 U
1 U/µl
$ 130.00 USD
109.00
130.00
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8
dsDNase
70600-202
1 000
2 U/µl
$ 1,055.00 USD
877.00
1055.00
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8
HL-dsDNase
70800-203
2500 U
5 U/µl
$ 2,365.00 USD
1961.00
2365.00
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9
Cod UNG
70500-202
1000 U
1 U/µl
$ 548.00 USD
456.00
548.00
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9
SAN High Quality ELISA Kit
70930-001
1x96 Well Compact Plate Format
$ 756.00 USD
655.00
756.00
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9
dsDNase
70600-203
2 500
5 U/µl
$ 2,365.00 USD
1961.00
2365.00
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9
HL-dsDNase
70800-150
50 kU
50 U/µl
$ 30,330.00 USD
26368.00
30330.00
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10
dsDNase
70600-150
50 kU
50 U/µl
$ 17,069.00 USD
14830.00
17069.00
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10
Cod UNG
70500-110
10 kU
1 U/µl
$ 3,920.00 USD
3472.00
3920.00
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10
HL-dsDNase
70800-110
100 kU
2 U/µl
$ 45,552.00 USD
39608.00
45552.00
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11
dsDNase
70600-110
100 kU
2 U/µl
$ 25,604.00 USD
22260.00
25604.00
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11
Cod UNG
70500-151
50 kU
1 U/µl
$ 14,128.00 USD
12281.00
14128.00
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11
HL-dsDNase
70800-120
500 kU
2 U/µl
$ 170,682.00 USD
148412.00
170682.00
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12
Cod UNG
70500-150
50 kU
50 U/µl
$ 14,128.00 USD
12281.00
14128.00
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12
dsDNase Glycerol Free
70610-50
5000 U
80 - 140 U/µl
$ 4,031.00 USD
3505.00
4031.00
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12
HL-dsDNase Glycerol-free
70810-50
5000 U
80 - 140 U/µl
$ 4,031.00 USD
3505.00
4031.00
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13
Cod UNG Glycerol Free
70510-105
5000 U
50 U/µl
$ 2,598.00 USD
2256.00
2598.00
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14
Cod UNG Glycerol Free
70510-150
50 kU
50 U/µl
$ 14,128.00 USD
12281.00
14128.00
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Note: Article no. 71502-202 and 71502-120 are provided with IsoPol® BST+ 10X Buffer and MgSO4 .
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Bioprocessing with Salt Active Nucleases  - High Salt Conditions
SAN HQ
now available in
GMP grade