From: Application of phage display to high throughput antibody generation and characterization
Step | Number | Percentage |
---|---|---|
(a) Mammalian expression of receptor extra-cellular domains | Â | Â |
Sequenced confirmed clones | 157 | 100% |
Detected in supernatant by western blot | 108 | 69% |
50 μg purified product generated from 50 ml culture | 67 | 62% (43% cumulative) |
(b) Success rate of antigens in generation of antibodies | Â | Â |
Mammalian antigens for selection | 205 | 100% |
Antigens generating antibody | 181 | 88% |
Bacterial antigens for selection | 192 | 100% |
Antigens generating antibody | 103 | 54% |
Total antigens for selection | 404 | 100% |
Antigens generating antibody | 292 | 72% |
(c) Success rate for primary antibody screening | Â | Â |
Number of antibodies screened | 38,164 | 100% |
Number positive on primary screen | 9,384 | 24.6% |
Unique clones generated | 7,236 | 77.1% (19% cumulative) |
(d) Success rate for secondary antibody screening | Â | Â |
Clones undergoing secondary specificity screening | 4,437 | 100% |
Positive in secondary screening | 3,400 | 76.6% |
Specificity demonstrated | 2,725 | 80.1% (61% cumulative) |
(e) Success rate in immunohistochemistry | Â | Â |
Stained on tissue microarrays | 381 | 100% |
Positive staining observed | 143 | 37% |