0 item(s)
View cart
You have no items in your shopping cart.
For Research Use Only (RUO)
$876.00
Sku: 1301302
Product Description
A sandwich-format solid-phase ELISA for quantitation of residual host cell proteins (HCPs) derived from E.coli K-12 strains (e.g., DH5α, Top10, JM109) in plasmid DNA (pDNA) intermediates or bulk prepared by alkaline lysis. The kit uses an affinity-purified capture antibody (sheep polyclonal) raised against alkaline-lysis-derived E.coli K-12 HCPs to capture HCPs, a biotinylated detection antibody and streptavidin–HRP for signal generation. Detection is performed with TMB substrate (read at 450 nm after stop), and sample concentrations are calculated against a six-point calibration curve using a 4-parameter logistic (4-PL) regression. The kit is supplied as 96 tests (8 × 12 strips) and is intended for research use only.
Technical Specifications
| Parameter | Details |
|---|---|
| Assay format | 96-well microtiter strips (8 × 12 strips), precoated with sheep anti-E.coli HCP-AC capture antibody |
| Intended sample types | pDNA intermediates, in-process samples, drug substance and drug product prepared by alkaline lysis |
| Linearity & Range | 3 - 729 ng/mL (R² ≥ 0.990) |
| Lower Limit of Quantitation (LLOQ) | 3 ng/mL |
| Typical calibration model | 4-parameter logistic (4-PL) regression. Example parameters: A = 3.94571; B = -0.57096; C = 1732.85755; D = -0.05957; R² = 0.99973 |
| Specificity / Cross-reactivity | No cross-reactivity observed with CHO, Vero, HEK293T, and Hansenula polymorpha |
| Sample pH requirement | Sample pH should be between 6.5 and 8.5 |
| Typical incubation conditions | Sample incubation: 100 µL, 1 hour at RT with shaking (300–500 rpm) or 2 hours standing; Biotinylated antibody: 45 min at RT; Streptavidin-HRP: 15 min at RT; TMB: 15 min at RT |
| Kit size / Reactions | 96 tests/kit |
| Part / Product No. | Product No.: 1301302; Calibration Standard PNB002; Strips PNA002; other reagents PN* as labeled |
Features
- Alkaline-lysis‑targeted antibodies: Sheep polyclonal antibodies raised against HCPs derived from alkaline lysis of E.coli K-12, improving detection accuracy for alkalinely prepared pDNA samples.
- Sandwich ELISA sensitivity: Double-antibody sandwich format with biotin–streptavidin–HRP amplification enables a lower limit of quantitation of 3 ng/mL and a wide dynamic range up to 729 ng/mL.
- Ready-to-use precoated strips: Microtiter strips are pre-coated with capture antibody and supplied in sealed bags with desiccant for convenient setup and reduced plate-coating variability.
- Complete kit for routine testing: Includes calibration standard (lyophilized), reconstitution solution, wash buffer concentrate, biotinylated detection antibody, streptavidin‑HRP, TMB substrate and stop solution.
- Validated for common cell lines: Demonstrated no cross-reactivity with common non-E.coli host cells (CHO, Vero, HEK293T, Hansenula polymorpha).
Applications
- Quantitation of residual E.coli K-12 host cell proteins in plasmid DNA (pDNA) intermediates produced by alkaline lysis
- In-process monitoring of HCP clearance during pDNA purification
- Testing drug substance and drug product pDNA for residual E.coli HCPs (research use)
- Method validation support and spike-recovery experiments for pDNA analytics
Kit Contents
E.coli HCP-AC Calibration Standard (PNB002) — lyophilized (3 bottles)Store at 2-8 °C; reconstituted standard: 2-8 °C up to 30 days
Anti-E.coli HCP-AC Microtiter Strips (PNA002) — 8 well × 12 stripsStore sealed in bag with desiccant at 2-8 °C; opened strips: up to 60 days
Reconstitution Solution (PNC002) — 2 × 1.5 mLStore at 2-8 °C
Wash Buffer Concentrate (10×) (PNF001) — 3 × 25 mLStore at 2-8 °C; if crystalline, warm to 37 °C to dissolve before dilution
Anti-E.coli HCP-AC Biotinylated Conjugate (20×) (PNG002) — 1 × 600 μLStore at 2-8 °C; centrifuge briefly before use; prepare fresh 1× dilution before assay
Streptavidin-HRP (100×) (PNH002) — 1 × 140 μLStore at 2-8 °C; centrifuge briefly before use; prepare fresh 1× dilution before assay
TMB Substrate (PND002) — 1 × 12 mLKeep sealed, protected from light, store at 2-8 °C; equilibrate to RT 20 min before use; discard if colored
Stop Solution (PNI002) — 1 × 6 mL (1 M HCl)Store at 2-8 °C; corrosive — handle with appropriate PPE
Sealing Film (PNK001) — 3 piecesStore at 2-8 °C
* Total: 96 tests/Kit | Method: ELISA
Attention
• For Research Use Only (RUO). Not for clinical or diagnostic use.
• Intended for qualified technicians. Users should validate the assay for their sample matrix prior to routine use.
• Samples must be clear and free of insoluble material; centrifuge or filter before testing.
• Maintain sample pH between 6.5 and 8.5; samples outside this range may give abnormal results.
• Avoid repeated freeze–thaw cycles; for long-term storage, keep samples at −70 °C.
• Do not mix reagents from different lot numbers.
• Use ultra-pure water and ensure water temperature does not exceed 37 °C when preparing reagents.
• Wash wells thoroughly and avoid drying wells prior to substrate addition.
• TMB must be colorless before use; read plate within 30 minutes after adding stop solution.
• Stop solution is 1 M HCl — avoid contact with skin and eyes; use appropriate PPE.
• Avoid samples containing sodium azide (NaN3) because it inhibits HRP and will underestimate HCP levels.
• Prepare biotinylated antibody and streptavidin-HRP dilutions fresh for each assay; recommended single-use for some prepared solutions.
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 sample types are compatible with this kit?
The kit is optimized for pDNA samples and intermediates prepared by alkaline lysis from E.coli K-12 strains (e.g., DH5α, Top10, JM109). Samples should be clear and within pH 6.5–8.5.
Q2: What is the assay sensitivity and dynamic range?
The lower limit of quantitation (LLOQ) is 3 ng/mL and the assay is linear from 3 to 729 ng/mL (R² ≥ 0.990).
Q3: Can I incubate the plate without a thermoshaker?
Yes — if a microplate thermoshaker is not available, incubate samples standing for 2 hours at room temperature instead of 1 hour with shaking.
Q4: How should I handle reagents after opening?
Store opened components at 2–8 °C as specified (e.g., strips in bag with desiccant up to 60 days; reconstituted standard up to 30 days). Prepare certain dilutions fresh and use single-use when recommended.
Research Use Only
✔
Research Use Only (RUO). Not for clinical diagnostic use. – 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.
