|
Choice of Purification Method for high yields is dependent on the species. |
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| Sample |
Purification method |
Average Conc. (mg/mL)1 |
Mean ELISA Titer2 |
Recovery of Ag- specific Igs (%)3 |
Purity of Ig preparation (%)4 |
Stability at 4°C5 |
|
|
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| Pre-purification |
1.3 × 106 (1.9 × 105) |
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| Immune Rabbit |
CA-AS |
12.3 |
1.4 × 106 (3.6 × 105) |
100 |
> 95% |
NT |
| SEP-EASE (PEG) |
10.9 |
1.1 × 106 (9.6 × 104) |
85 |
> 95% |
NT |
|
| Protein A/G |
8.9 |
1.1 × 106 (6.3 × 104) |
84 |
> 95% |
NT |
|
| Protein G |
14.3 |
1.8 × 106 (2.0 × 104) |
100 |
> 95% |
NT |
|
|
|
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| Malaria-Exposed Human |
Pre-purification |
1.1 × 104 (962) |
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| CA-AS |
12.3 |
1.2 × 103 (176) |
11* |
> 95% |
Yes |
|
| SEP-EASE (PEG) |
14.5 |
1.6 × 104 (4 × 103) |
100 |
> 95% |
Yes |
|
| Protein A/G |
14.4 |
4.8 × 103 (95) |
44* |
>95% |
No |
|
| Protein G |
19.8 |
6.6 × 103 (2.1 × 103) |
60 |
> 95% |
No |
|
|
Data expressed as mean (± SEM) of three independent purification experiments. 1Protein concentration was determined by the absorbance of the solution at 280 nm with an extinction coefficient of 13.5 (standard for IgG) for a 1% IgG solution (10 mg/mL) 2Mean ELISA titer for an OD405 = 1, for rabbits tested against MSP1-p42(FVO) plate antigen, for humans tested against MSP1-p42(3D7) plate antigen 3Recovery of Ag-specific Igs is based on Mean ELISA titer. 4Determined by scanning densitometry on Coomassie Blue stained gels 5Samples were stored for 24 weeks at 4°C and then tested by SDS-gelelectrophoresis, NT = not tested *A statistically significant difference between the methods in the recovery of Ag-specific Ab by ANOVA. The individual groups were isolated with a Tukey's post test (p < 0.025). | ||||||
Bergmann-Leitner et al. Malaria Journal 2008 7:129 doi:10.1186/1475-2875-7-129 |
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