Advanced 2D HCP Analysis
We offer various additional services to enhance HCP analysis, providing a deeper understanding of HCP profiling and HCP antibody coverage. Each can be incorporated alone or in combinations to improve accuracy in HCP analysis. Customers can also request their own modification to better address their objectives.
In-depth analysis based on your objectives:
A. Quadrant Analysis
This method divides the 2D image into four quadrants, counting HCP and antibody spots to reveal coverage variations across molecular weight and PI. Take our in-house anti-HEK HCP antibody as an example:
The quadrant analysis shows almost 100% coverage in Q1 and Q2, and 93-94% coverage in Q3 and Q4. This information would be critical to the HEK antibody development.
Anti-HEK293 HCP Western Blot spot map
Anti-HEK293 HCP WB: Quadrant Coverage
Brown: Spots detected by anti-HEK293 HCP antibody
Blue: Spots not detected by anti-HEK293 HCP antibody
B. Exclusion of Drug Substance (DS) spots
Harvest or in-process samples contain various levels of DS, interfering with HCP profiling and HCP antibody coverage. We resolved this issue by locating, tagging, and removing all DS spots from 2D image. The report includes both the standard analysis with all spots and an advanced analysis removing DS spots. Please view here for more detailed info.
C. Exclusion of Negative Spots
Negative spots are those protein spots that bind to the secondary antibody in the absence of primary antibody. Their presence in HCP samples can lead to false positives in antibody coverage. To identify them, an additional 2D Western blot is performed using only the secondary antibody. Any detected negative spots are excluded from the HCP antibody coverage calculation.
If you have any questions on experiment design or additional analysis, feel free to contact our tech support team at support@appliedbiomics.com or send us an info request.