Host Cell Protein (HCP) by DIGE vs. ELISA

HCP Analysis: Advantages of 2D DIGE Western Blot Over ELISA & Standard 2D WB

	ELISA	Standard 2D Western Blot	2D DIGE Western Blot
Number of gels for each Ab		Two	One
Detection Method		Silver Staining	CyDye Labeling
HCP Composition and profiling	No
Detect protein modification	No
Detect protein degradation	No
Protein and WB in same gel	No	No
In-gel protein and WB comparison	No	No
Accuracy	Low	Low	High
	Inaccurate: In-direct reaction of antibodies with the antigen’s associated proteins	Protein spots are counted from the gel, NOT from the membrane	Transferred protein spots are scanned directly from the membrane
	False negative: 1) Coated Ab does not capture all HCP; 2) Surface denaturation of coating Ab can affect detection	Error caused by: 1) Gel to gel variation; 2) Gel to membrane variation	Accurate: Protein and Antibody signal on the same gel, thus avoiding any ambiguity
Consistency and Reproducibility	Lower	Lower	Higher