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This client is a Top 10 pharmaceutical company developing a product utilizing a lentiviral vector expressed in HEK 293 T cells.
Due to the complexity of the product, which contains proteins from both the virus, human cell line, and cell culture medium, the client used LC-MS to supplement their HEK293 HCP ELISA kit.
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Alphalyse helps biotech and pharmaceutical companies improve their understanding of the Host Cell Protein (HCP) in preclinical and clinical drugs. This knowledge enables better optimization of the downstream process for optimal removal of HCPs. If you would like to know more, visit our website
https://www.alphalyse.com/host-cell-protein-analysis/
If you would like to learn about our customer cases and follow our work within the research of Host Cell Proteins, you can follow us on LinkedIn: https://www.linkedin.com/company/alphalyse-inc./
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HEK 293 HCP ELISA Kit, 3G (F650S) – Cygnus Technologies
HEK 293 HCP ELISA Kit is used to monitor the optimal removal of HCP impurities as well as in routine final product release testing.
Source: www.cygnustechnologies.com
Date Published: 10/13/2021
View: 9635
HEK 293 HCP ELISA Kit, 3G Mammalian – cogershop.com
Expression of viral vectors in HEK 293 cells is a wely used procedure for obtaining sufficient quantities of a desired virus. The manufacturing and …
Source: www.cogershop.com
Date Published: 8/30/2021
View: 4395
Cygnus Technologies HEK 293 HCP ELISA KIT, 3G
HEK 293 HCP ELISA Kit, 3G; This kit replaces previous kit F650R; Storage:Standards -10°C to -30°C upon receipt, All other kit reagents 2°C to 8°C; CYGNUS …
Source: www.fishersci.com
Date Published: 1/7/2022
View: 5223
Human HEK 293 HCP ELISA | Easy 14 step protocol!
Human HEK 293 HCPs ELISA kit from Assay Genie is a high quality, simple protocol, ELISA kit to measure HEK 293 HCPs in serum, plasma, lysate samples.
Source: www.assaygenie.com
Date Published: 3/12/2022
View: 2454
HEK293T host cell protein ELISA kit – ENZ-KIT162
Sensitive measurement of HEK293T host cell protein · Low intra-assay variability allows for streamlined testing · High throughput format with results in 6 hours …
Source: www.enzolifesciences.com
Date Published: 1/29/2021
View: 6135
HEK 293 HCP ELISA Kit (F650S) – Szabo-Scandic
This HEK 293 HCP ELISA Kit, 3G, Cat. No. F650S, replaces the previous F650R kit while incorporating several improvements in performance.
Source: www.szabo-scandic.com
Date Published: 8/6/2022
View: 7820
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주제와 관련된 더 많은 사진을 참조하십시오 Can viral, bovine, and human proteins be quantified in one assay? | HEK293 HCP ELISA kit. 댓글에서 더 많은 관련 이미지를 보거나 필요한 경우 더 많은 관련 기사를 볼 수 있습니다.
주제에 대한 기사 평가 hek 293 hcp elisa kit
- Author: Alphalyse A/S
- Views: 조회수 83회
- Likes: 좋아요 없음
- Date Published: 2021. 10. 15.
- Video Url link: https://www.youtube.com/watch?v=UAEBfsF0CC8
HEK 293 HCP ELISA Kit, 3G
The absolute quantitation of HCP assays is exceedingly difficult for several reasons. First of all, there is no recognized reference preparation of HCP against which we can calibrate our assays. While ELISA is inherently a quantitative method when applied to a single analyte, ELISAs that attempt to measure simultaneously all of the hundreds of potential HCP impurities in the same well using a single reporter/detector system are at best semi-quantitative assays. Many arbitrary choices and assumptions are used when preparing anti-HCP antibodies and later the choice of material to use when preparing “standards”.
How to obtain the HCP material for immunogen is the first choice that must be made.
Do we use a null cell line or a mock transfected cell line or one with the actual product plasmid and what differences in HCP might we see from each?
At what point in the purification process do we obtain the HCPs for the immunogen or standards?
In almost every case the array and relative concentrations of HCPs in final product are different from the HCP in the ELISA standards.
How does one assign a total HCP concentration to an indeterminant mixture of many proteins with different molecular weights?
What effect will the different affinities of antibodies to various HCPs as well as the different concentrations of those antibodies have on the ability of the assay to be quantitative?
How do we know that we have antibody to all the HCPs present in a given sample type?
All of these questions and arbitrary choices mean HCP assays will at best be semi-quantitative methods capable of making only relative estimates of HCP concentrations from one sample to the next. We estimate the potential error in reporting an HCP level in a sample that has a different array of HCPs from the standards to be as high as 4-fold even if you have a very good broadly reactive antibody. The commonly expressed theoretical concern is that HCP assays might under-estimate results due to incomplete coverage of all HCPs. In reality, the quantitative errors in HCP assays due to the arbitrary choices and fundamental limitations of the method as discussed above can result in either over-estimation or under-estimation and tend to do so to about the same degree. In our experience, a well generated and affinity purified antibody will react to more than 70% of individual HCPs as demonstrated by traditional 2D Western blot correlated to silver stain. What is more important than the number of individual proteins with antibody reactivity, is the total mass of those proteins that are detected. In our experience and as you might expect from theoretical considerations, the proteins in highest concentration have the best chance of generating an antibody. As such, an antibody with about 70% reactivity to an individual HCP will react with those proteins that account for more than 95% of the total mass of HCP. Given this level of uncertainty in the absolute HCP concentration, the most important criteria by which we can judge any HCP assay is on other objective parameters by which any analytical method should be qualified or validated. Those criteria include specificity and accuracy as demonstrated by sample dilution linearity and spike recovery experiments, precision, robustness, and sensitivity. Provided the assay has sensitivity to detect at least a relative portion of the HCP in your final drug substance and the assay also meets the other objective analytical parameter specifications, such an assay is a valuable analytical method capable of demonstrating process control and reporting relative levels of HCP contamination from lot to lot as a release test.
Due to the impracticality of obtaining real final product HCPs that co-purify with product down to the final step, most of our HCP assays will utilize a source of HCPs from very upstream in the purification process. Those HCPs typically do not come from the actual product cell line but rather a null cell or mock transfected cell line. Once we have decided on the source material for the kit standards we first perform some partial purifications to remove non-HCP materials such as growth media additives, buffer salts and extraction reagents. Our initial approach to calibrate this processed material is to simply perform a BCA total protein assay using bovine serum albumin (BSA) as an arbitrary standard. When standards made with the HCP material calibrated by BCA give reasonable “stoichiometric agreement” with the amounts of antibody used in the ELISA then we feel the calibration by BCA is good enough. What is meant by “stoichiometric agreement” is that we know how much antibody is used in the ELISA. It is the quantity of antibody that actually dictates the analytical range and dose response curve of the ELISA. If we assume an average molecular weight for the total HCPs, then we can reasonably estimate HCP concentrations across the valid analytical range of the assay. When the BCA assay concentrations do not reasonably approximate the ELISA stoichiometry, we process the HCP material further. Such processing involves various purification steps to remove components registering in the BCA assay but that are in fact not HCP or at least not immunoreactive HCPs. This processing might also involve affinity purification against the anti-HCP antibody. The purification stops as soon as the BCA concentration gives a realistic stoichiometric agreement in the ELISA.
HEK 293 HCP ELISA Kit, 3G Mammalian
Le Kit F650S remplace le kit F650R Description Expression of viral vectors in HEK 293 cells is a widely used procedure for obtaining sufficient quantities of a desired virus. The manufacturing and purification process of these products leaves the potential for impurities by host cell proteins (HCPs) from HEK 293 cells. HCP impurities can result in adverse toxic or immunological reactions and thus it is desirable to reduce these impurities to the lowest levels practical. The polyclonal antibodies used in this kit have been generated against and affinity-purified using a mild lysate of HEK 293 cells to obtain HCPs typically encountered in initial product recovery steps. This kit can be used as a process development tool to monitor the optimal removal of host cell impurities as well as in routine final product release testing when comprehensively characterized using samples from your process. Antibody Affinity Extraction (AAE) was used to determine reactivity to individual HCPs in a mild lysate of HEK 293. AAE indicated reactivity to ~74% of the individual 2-D PAGE fractionated spots presumed to be HCP. This HEK 293 HCP ELISA Kit, 3G, Cat. No. F650S, replaces the previous F650R kit while incorporating several improvements in performance. The F650S kit uses a new broadly-reactive antibody exceeding the reactivity of the previous F650R antibody. In addition, F650S kit has increased sensitivity (LOD 0.2 ng/mL; LOQ 1 ng/mL) combined with a shorter Detection Ab and Sample/ Standard incubation time (1.5 h). Finally, the new sample and conjugate diluents were incorporated to enhance F650S stability. Product Information Product Type: ELISA Kit Storage: Standards -10°C to -30°C upon receipt, All other kit reagents 2°C to 8°C Target Expression System: HEK 293 Species Group: Mammalian Species: Human Fiche technique
Cygnus Technologies HEK 293 HCP ELISA KIT, 3G
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Human HEK 293 HCP ELISA (HUFI03434)
*Note: Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.
Before adding to wells, equilibrate the SABC working solution and TMB substrate for at least 30 min at 37°C. When diluting samples and reagents, they must be mixed completely and evenly. It is recommended to plot a standard curve for each test.
HEK293T host cell protein ELISA kit
Sensitive (30 ng/ml) ELISA for the quantitative determination of host cell protein contamination in bulk products expressed in E. coli expression systems.
ELISA, Colorimetric detection | Print as PDF
ENZ-KIT127-0001 96 wells Inquire for pricing
Do you need bulk/larger quantities?
Sensitive (10 ng/ml) ELISA for the quantitative determination of host cell protein contamination in bulk products expressed in Chinese Hamster Ovary (CHO) expression systems.
ELISA, Colorimetric detection | Print as PDF
ENZ-KIT128-0001 96 wells Inquire for pricing
Do you need bulk/larger quantities?
Quantitative protein standards for Western blotting internal controls or ELISA standards.
Purified from E. coli host cells., ELISA, WB | Print as PDF
ENZ-PRT121-0025 25 µg Inquire for pricing
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Quantitative protein standards for Western blotting internal controls or ELISA standards.
Purified from Chinese Hamster Ovary (CHO) host cells., ELISA, WB | Print as PDF
ENZ-PRT122-0025 25 µg Inquire for pricing
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WB | Print as PDF
ENZ-ABS260-0100 100 µg Inquire for pricing
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WB | Print as PDF
ENZ-ABS261-0100 100 µg Inquire for pricing
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WB | Print as PDF
ENZ-ABS262-0100 100 µg Inquire for pricing
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WB | Print as PDF
ENZ-ABS263-0100 100 µg Inquire for pricing
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WB | Print as PDF
ENZ-ABS264-1000 1 ml Inquire for pricing
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WB | Print as PDF
ENZ-ABS265-1000 1 ml Inquire for pricing
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Personal, Compact Plate Reader
ELISA, Colorimetric detection | Print as PDF
HEK 293 HCP ELISA Kit
This HEK 293 HCP ELISA Kit, 3G, Cat. No. F650S, replaces the previous F650R kit while incorporating several improvements in performance. Expression of viral vectors in HEK 293 cells is a widely used procedure for obtaining sufficient quantities of a desired virus. The manufacturing and purification process of these products leaves the potential for impurities by host cell proteins (HCPs) from HEK 293 cells. HCP impurities can result in adverse toxic or immunological reactions and thus it is desirable to reduce these impurities to the lowest levels practical. The polyclonal antibodies used in this kit have been generated against and affinity-purified using a mild lysate of HEK 293 cells to obtain HCPs typically encountered in initial product recovery steps. This kit can be used as a process development tool to monitor the optimal removal of host cell impurities as well as in routine final product release testing when comprehensively characterized using samples from your process. Antibody Affinity Extraction (AAE) was used to determine reactivity to individual HCPs in a mild lysate of HEK 293. AAE indicated reactivity to ~74% of the individual 2-D PAGE fractionated spots presumed to be HCP.
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