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Host Cell Protein (HCP) Assays

 

 

 

Click to view a complete list of Applied Biomics HCP assays

Applied Biomics Host Cell Protein (HCP) assays using 2D DIGE followed by fluorescent Western blot offer significant ADVANTAGES over the ELISA and standard 2D Western blot, with the following applications:

1) Characterizations of the HCP antibodies
2) Detecting HCP contaminants originated from cells that are used to manufacture therapeutic proteins and vaccines

Experimental Procedure: HCP 2D Western Blot (HEK293)
Example-1: CHO Cell HCP Assay
Example-2: E.coli HCP Assay
Example-3: E.coli High Mw HCP Assay

Advantages of DIGE over ELISA and Standard 2D in HCP Detection

Applied Biomics will perform HCP assays using HCP antibodies provided by customers. In addition, we cover the following HCP antibodies:

Anti-E.coli-HCP; Anti-Bacteria (Lactococcus lactis) HCP
Anti-Yeast (Saccharomyces cerevisiae) HCP; Anti-Yeast (Pichia pastoris) HCP
Anti-SF9 Insect Cell HCP; Anti-SF22 Insect Cell HCP
Anti-Vero Cell HCP; Anti-CHO-HCP; Anti-BHK-HCP; Anti-HEK 293-HCP; Anti-A549-HCP; Anti-MRC5-HCP; Anti-PER.C6-HCP
Anti-Bovine Serum Proteins; Anti-Mycobacterium Proteins; Anti-Goat-Milk Proteins

Experimental Procedure: HCP 2D Western Blot (HEK293)

 

 

 

protein labeling

Protein extraction
Protein assay
Protein labeling

 

HCP 2D gel

2D Gel image:
HEK293-Protein











Gel transfer to Membrane



HCP 2D Membrane


2D Membrane image:
HEK293-Protein



HCP 2D Membrane


2D Membrane image:
Anti-HEK293 HCP
WB Image



HCP image overlay


Image Overlay:
Green: HEK293-Protein
Red: Anti-HEK293 HCP


Spot Matching and Quantitation by DeCyder (100% matching, no ambiguity)


HEK293-HCP Protein Image

HCP Protein

Anti-HEK293 HCP WB Image

HCP Western Blot

Brown: 1949 reacting spots
Blue: 109 non-reacting spots (only present in protein image)
Ab Coverage of Anti-HEK293 HCP: 94.7% of 2058 spots from 293 HCP


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Example 1: Characterization of the CHO-HCP Antibody by 2D Western blot from the same 2D gel

 

CHO-Protein Image

CHO Protein

Anti-CHO HCP WB Image

Anti-CHO HCP Western Blot

Image Overlay: Green: CHO-HCP  Red: Anti-CHO HCP

CHO HCP Anti-CHO HCP image overlay

CHO-HCP Protein Image

CHO HCP Protein

Anti-CHO HCP WB Image

Anti-CHO HCP Western Blot

Brown: 2207 reacting spots
Blue: 89 non-reacting spots (only present in protein image)
Ab Coverage of Anti-CHO HCP: 96.1% of 2296 spots from CHO HCP

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Example 2: Characterization of E.coli-HCP antibodies from the same 2D gel

 

E.coli-Protein Image

E. Coli Protein

Anti-E.coli HCP WB Image

Anti-Ecoli HCP Western Blot

E. Coli-Protein Image

E.Coli protein

Anti-E. Coli HCP WB Image

Anti-Ecoli HCP Western Blot

Image Overlay: Green: E. Coli-HCP  Red: Anti-E.Coli HCP

E.coli HCP Anti-E. Coli HCP image overlay

E.coli-HCP Protein Image

E.Coli HCP Protein

Anti-E.coli HCP WB Image

Anti-E.Coli HCP Western Blot

Brown: 2220 reacting spots
Blue: 142 non-reacting spots (only present in protein image)
Ab Coverage of Anti-CHO HCP: 94% of 2362 spots from CHO HCP

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Example 3: Characterization of High-MW E.coli-HCP antibodies

 

E. Coli-Protein Image

E. Coli protein

Anti-E. Coli HCP WB Image

Anti-E. Coli HCP Western Blot

Image Overlay: Green: E. Coli-HCP  Red: Anti-E.Coli HCP

E. Coli-HCP Anti-E. Coli HCP image overlay

E.coli-HCP Protein Image

E. Coli-HCP protein

Anti-E.coli HCP WB Image

Anti-E. Coli HCP Western Blot

Brown: 1030 reacting spots
Blue: 3 non-reacting spots (only present in protein image)
Ab Coverage of Anti-E.coli high Mw HCP : 99.7% of 1033 High Mw spots from E.coli HCP.

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