Intermediates, semi-finished products and finished products of various biological products
Detection Method
Sandwich-ELISA
Intended Use
The Benzonase® kit is used for detection and quantitative determination of Benzonase® endonuclease in intermediates, semi-finished products and finished products of various biological products.
Contents of Kit
No.
Components
Size
Storage Conditions
1
Benzonase Microplate
96 well (12 strips of 8 wells)
2-8°C
2
Benzonase Standard
1×300 μL, 0.5 μg/mL
2-8°C
3
HRP-Conjugate Benzonase Antibody
15 mL
2-8°C
4
Wash Buffer 20×
30 mL
2-8°C
5
Sample Dilution
30 mL
2-8°C
6
Chromogen Solution A
8 mL
2-8°C
7
Chromogen Solution B
8 mL
2-8°C
8
Stop Solution
15 mL
2-8°C
Storage
1. Store all reagents at 2-8°C, and not be frozen or thawed. The product is valid for 12 months 2. All reagents must be brought to room temperature (20-25°C) prior to use.
Detection Range
0.05 - 5 ng/mL
Standard Curve
These standard curves are provided for demonstration only. A standard curve should be generated for each set of samples assayed.
Citations
Publication ()
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Background
Benzonase, also known as Serratia marcescens endonuclease, is a highly versatile enzyme that can degrade both single-stranded and double-stranded nucleic acids, including DNA and RNA. Unlike other nucleases, it does not have proteolytic activity, making it ideal for removing nucleic acid contaminants from protein samples and applications requiring complete digestion of nucleic acids. It is commonly used in bioprocessing to reduce the viscosity caused by genomic DNA. The enzyme functions optimally at a pH range of 8.0 to 9.2. It breaks internal phosphodiester bonds within nucleic acids, resulting in the generation of short oligonucleotides that are 3 to 8 bases in length. The active form of the enzyme is a dimer with two disulfide bonds in each monomer, which are crucial for its activity and stability.
The Benzonase Nuclease ELISA Assay Kit is a specialized tool used for the quantitative detection of residual nuclease content in intermediates, semi-finished products, and finished products of various biological products. Nucleases, including Benzonase, can pose a significant concern in the manufacturing and quality control of biological products, as they have the potential to degrade nucleic acids, which can affect the stability, efficacy, and safety of the final product. The kit provides a robust and standardized approach for assessing nuclease levels, enabling researchers to optimize manufacturing processes and implement quality control measures.
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References
Investigation of Enzymatic Behavior of Benzonase/Alkaline Phosphatase in the Digestion of Oligonucleotides and DNA by ESI-LC/MS
We have developed an ion-pairing HPLC?MS method that has sufficient separation power, selectivity, and sensitivity to investigate the enzymatic behavior of benzonase/alkaline phosphatase upon digestion of oligonucleotides and DNA. Mass spectrometry revealed that this enzyme pair can nonspecifically digest oligonucleotides and DNA into fragments ranging from 2 to 10 nucleotides, i.e., sizes suitable for routine mass spectrometric measurements. Trimers, tetramers, and pentamers are the most prominent digested products. This makes benzonase/alkaline phosphatase a promising choice for DNA and DNA adduct related studies that require a nonspecific enzyme. A computer software program developed in-house was critical in automating the processing of mass spectral data. The methodology described here provides a systematic approach for evaluating the behavior of DNA-cleaving enzymes by mass spectrometry.
Evaluation of Plasmid DNA Removal from Lentiviral Vectors by Benzonase Treatment
To improve the purity of lentiviral vector supernatants for clinical studies we have evaluated plasmid DNA removal from lentiviral vectors and also the extent of plasmid DNA associated with transduced CD34 cells in an ex vivo transduction protocol. Optimal conditions of plasmid DNA removal by benzonase treatment were established by varying the temperature, time, and benzonase concentrations in the reaction mix and were determined to be 50 units of benzonase per milliliter of vector supernatant at 37°C, for 15 min. No plasmid DNA was detected, suggesting efficient plasmid degradation was achieved under these experimental conditions. The infectious titer of benzonase-treated lentiviral vector (RRL-CMV-GFP) was nearly identical to the titer of untreated vector (2.3 ± 0.3 × 106 transduction units per milliliter (TU/ml) and 2.7 ± 0.3 × 106 TU/ml, respectively). Analysis of plasmid DNA in concentrated lentiviral vectors shows that concentration substantially decreases the amount of DNA per TU. Analysis of the extent of plasmid DNA associated with transduced CD34 cells in an ex vivo transduction protocol suggests that a minimal amount of plasmid is transferred to transduced cells if the vector supernatant was not previously treated with benzonase. In conclusion, benzonase treatment is effective in eliminating plasmid DNA from vector supernatants and treatment does not affect infectious titers. However, because there is minimal transfer of plasmid DNA to transduced cells under ex vivo transduction conditions, DNA removal from lentiviral vectors may not be essential for all ex vivo clinical applications.
COA
Publication 
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