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Product Description
SHENTEK® Residual 293T RNA Quantitation Kit is a one-step reverse transcription and fluorescence quantitative PCR (one-step RT-qPCR) assay designed to quantitate residual total RNA from 293T host cells in biopharmaceutical products at various manufacturing stages (in-process samples to final products). The kit uses specific primers and probes for 293T RNA and contains an Internal Positive Control (IPC) to monitor reaction performance and inhibition. The assay achieves femtogram-level sensitivity and is intended for research use. For nucleic acid extraction guidance, refer to SHENTEK® Residual Host Cell RNA Sample Preparation Kit (Product No. 1201205).
Technical Specifications
| Parameter | Details |
|---|---|
| Provided reactions | Reagents for 100 reactions |
| Sensitivity (claimed) | Detects residual 293T total RNA at femtogram (fg) level; lowest standard in recommended curve = 0.002 pg/µL (2 fg/µL) |
| Linear range / Standard curve | Recommended standard curve concentrations: 200 pg/µL (ST0) / Example curve used in protocol: ST1=20 pg/µL, ST2=2 pg/µL, ST3=0.2 pg/µL, ST4=0.02 pg/µL, ST5=0.002 pg/µL — effective linear range demonstrated from 2000 pg/µL (starting control dilution) down to 0.002 pg/µL depending on preparation |
| Reaction volume | 20 µL total reaction volume (15 µL qRT-PCR mix + 5 µL sample or control) |
| Sample input volume | 5 µL purified sample (or 5 µL standard/NTC/NCS) per 20 µL reaction |
| Thermal cycling conditions | Reverse transcription: 50 °C for 15:00 (1 cycle); Activation: 95 °C for 0:30 (1 cycle); 45 cycles of Denaturation 95 °C 0:10 and Annealing/Extension 60 °C 0:40 (fluorescence read at 60 °C) |
| Detection dyes / channels | 293T-RNA: FAM reporter; RNA-IPC: VIC reporter; Passive reference: ROX |
| IPC acceptance criteria | Ct-IPC of sample and NCS should be within ±1.0 Ct; significantly higher Ct-IPC in sample indicates possible inhibition. If IPC of NCS or sample fails, evaluate inhibition and recovery. |
| NTC / NCS criteria | Ct of NTC should be 2 cycles greater than mean Ct of the lowest standard; Ct of NCS should be larger than mean Ct of lowest standard and show VIC signal amplification |
| Plate layout / replicates | Recommended: at least five standard concentrations, NTC, NCS, IPC-NCS and IPC for each test sample; triplicate wells per sample/standard |
| Storage stability | Kit components stored at -20 °C; kit stable up to 24 months (check label expiry) |
| Compatible instruments | SHENTEK-96S, ABI 7500, Bio-Rad CFX96, Linegene 9600plus and other real-time PCR systems that support FAM/VIC/ROX |
| Units of report | Results reported as mean quantity in pg/µL (per instrument result export) |
Features
- One-step RT-qPCR: Reverse transcription and qPCR in a single-step workflow to reduce handling and contamination risk.
- High sensitivity: Analytical sensitivity at the femtogram level (standards include down to 0.002 pg/µL).
- Internal positive control (IPC): IPC primer/probe mix included to monitor reaction performance and detect sample inhibition.
- Pre-mixed primers & probes: 293T-specific primer & probe mix and RNA IPC primer & probe mix provided for convenience and consistency.
- Compatibility: Validated with multiple real-time PCR instruments (SHENTEK-96S, ABI 7500, CFX96, Linegene 9600plus).
- Ready for standard curve quantitation: Includes 293T RNA Control and recommended serial dilution scheme for absolute quantitation.
Applications
- Quantitation of residual 293T host cell total RNA in biopharmaceutical products
- Testing of in-process and final product samples for host cell RNA residues
- Method validation and sample recovery studies for residual host cell RNA
- Quality control during downstream processing and release testing (research use)
Kit Contents
293T RNA Control (NNA052) 50 µL × 1 tube-20 °C
One Step qPCR Buffer (NNB008) 500 µL × 2 tubes-20 °C, protect from light
One Step Enzyme MIX (NNC052) 100 µL × 1 tube-20 °C
293T RNA Primer & Probe MIX (NNC096) 400 µL × 1 tube-20 °C
RNA IPC Primer & Probe MIX (NNC053) 200 µL × 1 tube-20 °C
RNase-Free H2O (NND008) 1.2 mL × 3 tubes-20 °C
* Total: 100 reactions | Method: One-step RT-qPCR
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, clothing, gloves).
• Prevent RNase contamination: use DNase/RNase-free consumables, UV decontaminate work surfaces and tubes as described (30 min), disinfect with 75% ethanol.
• Thaw reagents at 2–8 °C or on ice; vortex and spin briefly before use.
• Protect light-sensitive reagents (One Step qPCR Buffer) from light.
• Treat samples with DNase prior to detection to remove genomic DNA influence; optimize DNase digestion conditions for sample type.
• Include at least five concentrations in the standard curve; run triplicates for standards and samples where possible.
• IPC criteria: Ct-IPC for sample and NCS should be within ±1.0 Ct; significantly higher Ct-IPC indicates inhibition.
• NTC and NCS acceptance criteria: NTC Ct should be ≥2 cycles above mean Ct of lowest standard; NCS Ct should be larger than mean Ct of lowest standard and show VIC signal.
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: How many reactions does one kit support?
This kit provides reagents sufficient for 100 reactions (as stated on the product packaging).
Q2: What is the recommended reaction setup and sample input volume?
Use a 20 µL total reaction volume (15 µL qRT-PCR mix + 5 µL sample or control). Prepare qRT-PCR mix according to the protocol.
Q3: What are the thermal cycling conditions?
Reverse transcription 50 °C for 15:00, Activation 95 °C for 0:30, then 45 cycles of 95 °C 0:10 (denaturation) and 60 °C 0:40 (anneal/extend; read fluorescence at 60 °C).
Q4: What should I do if IPC indicates inhibition?
If sample Ct-IPC is significantly higher than IPC-NCS (greater than ±1.0 Ct difference), investigate inhibitors in the sample, consider sample dilution, re-extraction, or additional purification; prioritize recovery results if spike-in controls were run.
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.
