KRIBIOLISA E.coli HCP (Generic Coverage) ELISA

Name: KRIBIOLISA E.coli HCP (Generic Coverage) ELISA
SKU: KBBP01
Availability: InStock

SKU: KBBP01

Rs 26,200.00

Immunoassay for the quantitative measurement of E.coli Host Cell Proteins as a contaminant in various biological preparations.

Availability: 4 – 5 days | Pack Size: 1 x 96 wells

Introduction:
A variety of proteins and pDNA which are used as therapeutic agents in humans and animals are produced through recombinant expression in E.coli. The manufacturing and purification process of these products tends to leave the potential for contamination by Host Cell Proteins (HCPs) from E.coli which may result in adverse toxic or immunological reactions that ultimately affect the efficacy of the therapeutic agent. The simple, objective and semi-quantitative ELISA is a highly sensitive method that aids in purification process development, process control, quality control and product release testing optimally.

Intended Use:
This generic kit is intended in determining the presence of E.coli Host Cell Proteins contamination in various products that are manufactured through recombinant expression in E.coli. The kit has been validated successfully for testing of final and in process product HCPs in variety of products regardless of growth and purification process.

Principle:
This assay is based on the Sandwich ELISA procedure. Samples containing E-coli HCPs are reacted with antiE.coli:HRP antibody simultaneously in the microtiter wells already coated with affinity purified capture anti-E.coli antibody. This immunological reaction results in formation of a sandwich complex of solid phase antibody-HCPenzyme labeled antibody. The wells are washed to remove any unbound reactants as per the wash procedure (see Assay procedure section mentioned below). The substrate Tetramethyl Benzidine is then reacted. The amount of hydrolyzed substrate is read on a microtiter plate reader and it is directly proportional to the concentration of E.coli HCPs present.

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Additional information

Pack Type	ELISA 96 wells
Species Reactivity	Ecol cells
Sample Type	Cell Culture Supernatant and other biological samples
Assay Range	3.125 – 200 ng/ml
Sensitivity	1.875 ng/ml
Interference	Preparations of the known factors were assayed for interference. No significant interference was observed.
Specificity	The assay recognises host cell proteins including low and high molecular weight proteins.
Principle Of Assay	Double Antibody, Sandwich ELISA
Detection Method	Absorbance measured at 450nm, Colorimetric
Conjugate-Enzyme Reaction	Uses HRP enzyme conjugate with TMB chromogenic substrate for color development
Quality Certification	Validated as per KRISHGEN's Gold Ring 7-Point Program
KRISHGEN Mandate	High Performance Assays at Affordable Prices
Regulatory Status	Research Use Only.
Shipping Temperature	Room Temperature / 2-8 degrees Celsius
Storage Temperature	if all the wells in the kit are not being used, store the unused wells in the foil pouch containing the desiccant pack, The kit and its components are to be stored as indicated in the IFU (instructions for use). To ensure quality of your results, well sealed and store at the indicated temperature.
ELISA Type	Double Antibody, Sandich ELISA
ELISA Interpretation	on request, Quantitative Results Intepretation (% Binding/OD450), Software (MS Excel) available, to support interpretation of results.
Shelf Life Available	8 -10 months at time of shipping
Alternate Names / Synonyms	E.coli HCP; ecol HCP; host cell proteins
Research Area	Biosimilars; Monoclonal Antibodies; small molecules; therapeutic drug monitoring assays
Disclaimer	The data indicated herein with specifications are changed from time to time at time of production. Please refer the most current IFU (instructions for use) accompanying the kit.

Protocol

Assay Procedure:
It is strongly recommended that all Standards and Samples be run in duplicates or triplicates. A standard curve is required for each assay.
1. Add 100 ul prepared Standards and diluted Samples to respective wells.
2. Cover the plate with a sealer and incubate for 90 minutes at 37 Degree Celcius.
3. Aspirate and wash plate 4 times with diluted Wash Buffer (1X) and blot residual buffer by firmly tapping plate upside down on absorbent paper. Wipe of any liquid from the bottom outside of the microtiter wells as any residue can interfere in the reading step.
4. Pipette 100 ul Biotinylated E.coli HCP Antibody Working Solution to all wells.
5. Cover the plate with a sealer and incubate for 60 minutes at 37 Degree Celcius.
6. Aspirate and wash as per Step (4) above.
7. Pipette 100 ul Streptavidin:HRP Conjugate Working Solution to all wells. Mix well.
8. Cover the plate with a sealer and incubate for 30 minutes at 37 Degree Celcius.
9. Aspirate and wash as per Step (4) above.
10. Pipette 100 ul TMB Substrate in all the wells.
11. Incubate the plate at 37 Degree Celcius for 20 minutes. DO NOT SHAKE or else it may result in higher backgrounds and worse precision. Positive wells should turn bluish in color.
12. Pipette 100 ul of Stop Solution to all wells. The wells should turn from blue to yellow in color.
13. Read the absorbance at 450 nm with a microplate within 10-15 minutes after addition of Stop solution.