AccuSignal™ Nuclease ELISA Kit for Impurity Detection

Produktnummer ABIN7505815

Produktdetails

Highlights:

• Nachweis von Verunreinigungen bei der Herstellung von biologischen Arzneimitteln.
• Der AccuSignal™ Nuclease ELISA Kit ist für die empfindliche und zuverlässige Quantifizierung von Nukleasen in therapeutischen Produkten, einschließlich DENARASE®, Benzonase® und Turbonuclease, konzipiert.
• Bietet ein breites Quantifizierungsspektrum bei hervorragender Verdünnungslinearität, was das Vertrauen in die Genauigkeit der Ergebnisse über ein breites Spektrum von Nukleasekonzentrationen stärkt.
• Liefert konsistente und wiederholbare Ergebnisse, die sich durch minimale Variabilität innerhalb und zwischen den Tests auszeichnen.
• Maßgeschneiderte Antikörperspezifität ermöglicht die Anwendung auf verschiedene Materialien, wobei verschiedene Nukleaseprodukte von mehreren Anbietern verwendet werden können.
• Bestandteile: 96-Well-Streifenplatte, Plattenversiegler, Nuklease-Standard, biotinylierter Nachweisantikörper, Streptavidin-HRO (100X), Puffer.

Target: Nuclease

Nachweismethode: Colorimetric

Wirt: Kaninchen

Klonalität: Polyklonal

Methodentyp: Sandwich ELISA

Detektionsbereich: 0.03 ng/mL - 20 ng/mL

Untere Nachweisgrenze: 0.03 ng/mL

Applikation: ELISA, Impurity Detection (Imp De)

Verwendungszweck: Nuclease ELISA kit is designed for the quantitative detection of Nuclease/NucA in serum, plasma, and hybridoma cell supernatants.

Marke: AccuSignal™

Analytische Methode: Quantitative

Spezifität: Benzonase, Nuclease, Endonuclease, Serratia marcescens endonuclease, NucA, Denarase

Sensitivität: 3 ng/mL

Bestandteile: This kit contains: Nuclease antibody coated 96-well strip plate, Plate Sealer, Nuclease standard, biotinylated Detection Antibody, streptavidin-peroxidase conjugate, along with buffers and protocol.

Benötigtes Material:

• Microplate shaker (up tp 450 rpm)
• Interval timer
• Multichannel pipettor (50-300 µL)
• Precision single pipettes (10 µL, 35 µL, 100 µL, 1000 µL, etc.)
• Disposable pipette tips
• Deionized water
• Disposable microcentrifuge Tube(s) or microplate
• Polypropylene centrifuge tubes (15 mL)
• Spectrophotometer microplate reader (450 nm absorbance, 630–650 nm reference filter)
• Disposable gloves
• Graduated cylinder
• Reagent reservoirs
• Vortex mixer
• Stir plate & magnetic stir bar
• Absorbent paper

Produktspezifische Information:

Das Nuclease ELISA Kit ist ein speziell entwickeltes, unverzichtbares Werkzeug, das die Menge an Nukleasen in therapeutischen Produkten, einschließlich DENARASE®, Benzonase® und Turbonuclease, sensibel und robust quantifiziert. Dieses Kit verwendet ein Sandwich-ELISA-Format, das die Spezifität und Affinität von vorimmobilisierten Anti-Nuklease-Antikörpern nutzt. Die biotinkonjugierte Erkennung in Kombination mit Streptavidin-HRP garantiert eine beispiellose Empfindlichkeit und ermöglicht eine genaue Messung von Nukleasen in verschiedenen therapeutischen Materialien. Es erleichtert die Bewertung des Nukleasegehalts in biologisch gewonnenen Therapeutika und sichert deren Stabilität und Qualität während des Herstellungsprozesses. Zusätzlich hilft es bei der Bewertung von Nukleasen in Vektoren für Gentherapien, einem kritischen Faktor für die Aufrechterhaltung der Integrität dieser Vektoren für eine effektive therapeutische Genlieferung. Es unterstützt auch bei der Validierung der Nukleasegehalte in pharmazeutischen Formulierungen, schützt deren Wirksamkeit und verhindert mögliche Zersetzung. Indem es eine zuverlässige und standardisierte Methode zur Quantifizierung von Nukleasen bietet, stellt das Nuclease ELISA Kit nicht nur die Einhaltung strenger regulatorischer Anforderungen sicher, sondern beschleunigt auch Forschungs- und Entwicklungsprozesse.

Hauptanwendungsgebiete für das AccuSignal™ Nuclease ELISA Kit

Biologische Therapeutika: Bewertung des Nukleasegehalts in biologisch gewonnenen Therapeutika, um deren Stabilität und Qualität während der Produktion zu garantieren.

Gentherapie-Vektoren: Unterstützung bei der Beurteilung von Nukleasepräsenz in Vektoren für Gentherapien, wesentlich für die Erhaltung ihrer Integrität für erfolgreiche Genlieferungsbehandlungen.

Impfstoffproduktion: Bestätigung der Nukleasekonzentrationen in pharmazeutischen Formulierungen, um deren Wirksamkeit zu gewährleisten und potenzielle Zersetzung zu verhindern.

Der benutzerfreundliche Ansatz und die robuste Methodik vereinfachen Arbeitsabläufe und ermöglichen es Wissenschaftlern, sich mit Vertrauen auf das Vorantreiben therapeutischer Innovationen zu konzentrieren. Als unverzichtbares Instrument fördert es Präzision, Zuverlässigkeit und Effizienz bei der Bewertung von Nukleasen in therapeutischen Produkten und gewährleistet gleichzeitig Sicherheit und Wirksamkeit.

Anwendungsinformationen

Applikationshinweise: This kit was shown to positively detect commercially available nuclease variants, such as Benzonase, Denarase and other analogs.

Testdauer: 3 h

Aufbereitung der Reagenzien:

Preparation of Standards & Test Samples

1. Reconstitute the standard vial with 1.0 mL deionized water to obtain a final concentration of 200 ng/mL.
Note: This is the “reference stock solution” that will be used below to make the standards.
2. Prepare dilutions of standard.
3. Test samples should be diluted in Nuclease Kit Sample Buffer based on empirically determined criteria for each sample.

Detection antibody working solution preparation

1. Add 110 µL of conjugated antibody to 11 mL of Sample Buffer for use in a full 96-well assay.
2. Mix well by pipette or inversion. Do not vortex.
3. Distribute antibody working solution as described in the assay procedure.

Note: Volumes may be adjusted so long as final working concentration remains as specified.

Streptavidin-HRP Working Solution Preparation

1. Add 110 µL of Streptavidin-HRP to 11 mL of Sample Buffer, respectively for use in a full 96-well assay.
2. Mix well by pipette or inversion. Do not vortex.
3. Distribute Streptavidin-HRP working solution as described in the assay procedure.

Note: Volumes may be adjusted so long as final working concentration remains as specified

Wash Buffer (1X) Preparation

1. Add 50 mL of Nuclease Kit Wash Buffer (10X) to 450 mL of deionized water.
2. Mix for at least 10-minutes using a magnetic stir bar.

Note: The wash buffer (1X) can be stored at room temperature (15°C to 25°C) for up to 2-weeks, after which it should be discarded.

Testdurchführung:

1. To each well add 100 µL of unknown or standard sample per well and incubate at room temperature for 60-minutes with shaking at 450 revolutions per minutes (rpm) on a shaker.
2. Wash the wells with Wash Buffer as follows:
• Decant the contents of the wells manually with a hard, rapid downward motion. Fluid should be captured in a receptable designed to collect the waste. Remove all residual reagent from the microplate by tapping it on absorbent paper with the opening facing downwards.
• Fill each well with 300 µL of Washing Buffer with a multichannel pipettor.
• Decant the Washing Buffer from the wells with a hard, rapid downward motion. Remove all residual solution from the microplate by tapping it on absorbent paper with the opening facing downwards.
• Repeat steps ii and iii two more times (total of 3 washings). Do not leave any residual moisture in the wells on each washing step.
3. To each well add 100 µL of detection antibody working solution
4. Incubate at room temperature for 60-minutes, covered to protect from light, with shaking at 450 rpm.
5. Following 60-minute incubation, wash with Wash Buffer as follows:
• Decant the contents of the wells manually with a hard, rapid downward motion. Fluid should be captured in a receptable designed to collect the waste. Remove all residual reagent from the microplate by tapping it on absorbent paper with the opening facing downwards.
• Fill each well with 300 µL of Washing Buffer with a multichannel pipettor.
• Decant the Washing Buffer from the wells with a hard, rapid downward motion. Remove all residual solution from the microplate by tapping it on absorbent paper with the opening facing downwards.
• Repeat steps ii and iii two more times (total of 3 washings). Do not leave any residual moisture in the wells on each washing step.
6. To each well add 100 µL of Streptavidin-HRP working solution
7. Incubate at room temperature for 20-minutes, covered to protect from light, with shaking at 450 rpm.
8. Following 20-minute incubation, wash with Wash Buffer as follows:
• Decant the contents of the wells manually with a hard, rapid downward motion. Fluid should be captured in a receptable designed to collect the waste. Remove all residual reagent from the microplate by tapping it on absorbent paper with the opening facing downwards.
• Fill each well with 300 µL of Washing Buffer with a multichannel pipettor.
• Decant the Washing Buffer from the wells with a hard, rapid downward motion. Remove all residual solution from the microplate by tapping it on absorbent paper with the opening facing downwards
• Repeat steps ii and iii two more times (total of 3 washings). Do not leave any residual moisture in the wells on each washing step.
9. Next add 100 µL per well of room temperature TMB solution and incubate the plate with TMB solution at room temperature for 20 minutes (covered and protected from light).
10. Then add 100 µL of stop solution per well. Gently tap the plate to mix, ensuring no bubbles are formed, and read plate within 5 minutes after stopping the reaction.
11. On plate reader, measure absorbance at 450 nm with the reference wavelength set at 630–650 nm.

Ergebnisberechnung:

Follow the steps below to estimate the nuclease concentration of the test samples.

1. Calculate the relative OD 450 using the following formula: Relative OD 450 = (OD 450 of well) – (OD 630-650 nm of the well)
2. Calculate the mean relative OD 450 of the replicates for each standard solution.
3. Plot the standard solutions data as mean relative OD 450 for each standard solution (Y) vs the respective concentration of the standard solutions (X).
4. Fit the standard solution data with a 4-parameter logistic (4-PL) curve. Weight by 1/Y^2 is intended to be used during generation of 4-PL curve
5. Estimate the Nuclease concentration of each test sample well using interpolation from the standard curve. Calculate the average of each respective sample solution concentration.

Note: If the assay samples are from dilutions, multiply the concentrations obtained from interpolations by the dilution factor.
Note: If the spectrometer used for the assay does not automatically subtract the reference wavelength, do this manually.

Testpräzision: Intra-and inter-assay CV% <20%

Beschränkungen: Nur für Forschungszwecke einsetzbar

Handhabung

Lagerung: 4 °C

Informationen zur Lagerung: The kit and reagents should be stored at 2-8°C. Allow reagents to reach room temperature (18-26°C) before use and may be used until the expiration date. It is recommended to aliquot the reconstituted standard solution and store it at -20°C to avoid freeze/thaw cycles.

Antigendetails

Target: Nuclease

Andere Bezeichnung: nucA (Nuclease Produkte)

Synonyme: F15D2.37 Kit, F15D2_37 Kit, 5'-3' exonuclease family protein Kit, Nuclease Kit, AT1G29630 Kit, Nuc Kit

Hintergrund: Nucleases are secreted by Serratia marcescens into the medium it surrounds. The enzyme is a sugar-nonspecific hydrolase, capable of cleaving both RNA and DNA in either double or single stranded form. It requires divalent cations, preferably Mg2+, and is functional across a broad pH range from 6 to 10 (optimal at 8-8.5) and wide temperature ranges between 35°C and 44°C. The ability of Serratia to secrete nuclease appears to be regulated. Bacterial cultures at differing cell densities display different kinetics and efficiencies of nuclease secretion [i.e. growth medium, growth conditions, and host cell mutations]. Anti-Nuclease/NucA Antibody is useful for researcher interested in identifying nucleic acid contamination.