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Product Description
SHENTEK® Residual E1B DNA Quantitation Kit is designed for absolute quantitation of residual E1B DNA derived from host cells (e.g., HEK293 and derivative cell lines such as 293T and 293F) in various stages of biopharmaceutical manufacturing (from in-process samples to final products). The kit uses a fluorescent quantitative PCR (qPCR) assay with an internal positive control (IPC) to monitor PCR performance and includes an E1B linear DNA Control for preparation of standard curves and for detection of linearized samples. The kit is supplied for 100 reactions and is intended for research use only. For sample extraction procedures, refer to the SHENTEK® Residual Host Cell DNA Sample Preparation Kit (Product No. 1104191).
Technical Specifications
| Parameter | Details |
|---|---|
| Intended reactions per kit | Reagents for 100 reactions |
| Detection format | Absolute quantitation by fluorescent qPCR (FAM-labeled target, VIC-labeled IPC, ROX passive reference) |
| Standard curve concentration range (prepared) | ST1 to ST6: 4.34 × 10^6 to 4.34 × 10^1 copies/μL (prepared from E1B linear DNA Control; original ST = 4.34 × 10^8 copies/μL before serial dilution) |
| Reaction volume | 30 μL total reaction volume (20 μL qPCR MIX + 10 μL sample or control) |
| qPCR MIX composition (per reaction) | qPCR Reaction Buffer 15.9 μL, E1B Primer&Probe MIX 2.8 μL, IPC MIX 1.3 μL (total qPCR MIX = 20 μL) |
| Thermal cycling | Activation: 95 °C 10:00; Denaturation: 95 °C 0:15; Annealing/Extension: 60 °C 1:00; 40 cycles; fluorescence read during annealing/extension |
| Instrument settings / analysis | Manual Ct with threshold = 0.02 for both E1B-DNA and IPC; Automatic baseline; Standard curve (absolute quantitation) analysis |
| Negative control acceptance | Ct of NTC should be ≥ 35.00 cycles or undetermined |
| IPC acceptance criteria | Ct(IPC) of sample should be within ±1.0 Ct of Ct(IPC) of NCS; a significantly higher Ct(IPC) indicates potential PCR inhibition |
| Sample input for preparation | 100 μL test sample (or 100 μL DDB for negative control sample preparation prior to extraction) |
| Shelf life | Kit components can be stored under stated conditions for up to 24 months; check label expiration date |
| Limit of quantification / limit of detection | Not numerically specified in the manual. Users should establish LOD/LOQ by validation; standard curve lowest point in user protocol is 4.34 × 10^1 copies/μL |
Features
- High specificity and sensitivity: Fluorescent qPCR assay with sequence-specific primers and probe for E1B enabling specific detection of residual E1B DNA.
- Internal positive control (IPC): Included IPC MIX monitors PCR performance and helps identify sample inhibition.
- Ready-to-use components: Pre-formulated qPCR Reaction Buffer, Primer & Probe MIX and IPC MIX minimize setup errors and speed assay setup.
- E1B linear DNA Control: Supplied lyophilized linear DNA control for preparation of serial dilutions and construction of an absolute-quantitation standard curve.
- Compatibility: Validated on multiple real-time PCR instruments (e.g., SHENTEK-96S, ABI 7500, Bio-Rad CFX96, qTOWER3G, LightCycler480).
- For residual host cell DNA testing: Designed specifically to quantitate residual E1B DNA from HEK293 and derivative cell lines in biopharmaceutical samples.
Applications
- Quantitation of residual E1B DNA in biopharmaceutical products (in-process and final products)
- Quality control and release testing for products produced in HEK293 and derivative cell lines (e.g., 293T, 293F)
- Method validation and assay development for residual host cell DNA analysis
- Assessment of DNA clearance and downstream process effectiveness
Kit Contents
E1B linear DNA Control (lyophilized powder × 1 tube) — Part No. NNA014-20 °C
qPCR Reaction Buffer (850 μL × 2 tubes) — Part No. NNB002-20 °C, protect from light
E1B Primer & Probe MIX (300 μL × 1 tube) — Part No. NNC034-20 °C, protect from light
IPC MIX (150 μL × 1 tube) — Part No. NNC066-20 °C, protect from light
DNA Dilution Buffer (DDB) (1.5 mL × 3 tubes) — Part No. NND001-20 °C
* Total: 100 reactions | Method: qPCR (fluorescent quantitative PCR, absolute quantitation)
Attention
• For research use only (RUO). Not for diagnostic or therapeutic use.
• Read Material Safety Data Sheets (MSDS) and follow handling instructions.
• Wear appropriate PPE (protective eyewear, mask, lab coat, gloves).
• Use DNase-free consumables and low-retention tips to avoid contamination and loss.
• Thaw reagents at 2–8 °C or on ice; vortex and briefly centrifuge before use.
• Protect light-sensitive reagents (qPCR Reaction Buffer, Primer&Probe MIX, IPC MIX) from light.
• Irradiate work surfaces, pipettes and tubes with UV for 30 minutes and disinfect with 75% ethanol prior to setup to reduce contamination risk.
• Ensure lyophilized E1B linear DNA Control is fully dissolved (add 55 μL ddH2O and mix as instructed).
• Include at least five standard concentrations and run samples in triplicate for reliable quantitation.
• NTC Ct should be ≥ 35 or undetermined; check IPC Ct to assess inhibition (sample IPC Ct must be within ±1.0 Ct of NCS IPC Ct).
• Establish assay-specific LOD/LOQ during method validation; lowest manual standard = 4.34 × 10^1 copies/μL.
Quality Management & Certifications
Quality System
Download QMSQMS (ISO, GMP)
Download CertificateQuality Advantages
View ReportQuality Control Process
Download ProcessTechnical Resources & Downloads
Frequently Asked Questions (FAQ)
Q1: What is the detection method used by this kit?
The kit uses fluorescent quantitative PCR (qPCR) for absolute quantitation of E1B DNA (FAM-labeled target, VIC-labeled IPC, ROX passive reference).
Q2: How many reactions are provided per kit and what is the recommended reaction volume?
The kit provides reagents for 100 reactions. The recommended total reaction volume is 30 μL (20 μL qPCR MIX + 10 μL sample/control).
Q3: What is the standard curve range included in the user protocol and is LOQ specified?
The protocol prepares a standard curve with concentrations from 4.34 × 10^6 down to 4.34 × 10^1 copies/μL (ST1–ST6). A numeric LOQ/LOD is not specified in the manual; users should determine assay-specific LOD/LOQ during validation.
Q4: How should I interpret IPC and controls?
IPC (internal positive control) monitors PCR performance and sample inhibition. The Ct(IPC) of a sample should be within ±1.0 Ct of the Ct(IPC) of the NCS. The NTC should have Ct ≥ 35 or be undetermined. A significantly higher Ct(IPC) in a sample indicates possible inhibition; consider dilution or purification.
Research Use Only
✔
Research Use Only (RUO) – This product is intended for laboratory research purposes only and not for clinical or regulatory diagnostic use.
✔
Not intended for use in USDA or FDA regulated diagnostic testing or official compliance testing.
When can I expect my order to ship?
Most orders are filled and shipped within 2-3 business days from the time they are received.
Our standard shipping usually take 2-5 days.
We also provide express shippping for time-sensitive deliveries.
Email contact@biofargo.com if you have any requirements.
