method
A technology of immunoglobulin and solution, which is applied in the improvement of purified IgG and the improvement of purified IgG, and can solve problems such as being infected by HIV
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Embodiment 1
[0070] (a) According to the prior art method of Steinbuch and Audran ((1969) Arch. Biochem. Biophys. 134, 279-294).
[0071] A portion of the frozen NA pellet (PPT) was resuspended in sufficient sodium acetate buffer and the NA PPT suspension was stirred for several hours until most of the IgG dissolved at ambient temperature while keeping the pH constant at 4.8.
[0072] Defatting was performed by adding octanoic acid (OA) to the suspension and subsequent incubation for 240 minutes. Calcium phosphate was then added and the suspension was stirred for an additional 60 to 90 minutes. Precipitated proteins, lipids or lipoprotein complexes and other contaminants precipitated are removed by filtration under these conditions.
[0073] The OA-filtrate was subjected to ultrafiltration to a protein concentration of 3%, followed by diafiltration against water for injection (WFI). The pH was continuously switched to pH 4.1 with 0.2 mol / L HCl during diafiltration. After diafiltration t...
Embodiment 2
[0093] This example demonstrates the effect of different pH adjustments using Tris buffer on IgG yield compared to the existing pH shift of 6.5.
[0094]Starting from the PPT-NA described in Example 1, a low pH 4 incubated solution was generated as described above. Eight aliquots (1 kg each) were extracted and adjusted to pH 5.2, 5.4, 5.6, 5.8, 6.0, 6.2, 6.4 with 1M Tris buffer. The pH of the last aliquot was adjusted using 0.2M NaOH according to the prior art method. pH adjustments were made over a period of 90 minutes, followed by 90 minutes of incubation at ambient temperature as described above. The precipitate formed was then removed by filtration. The filtrate was then loaded onto an AIEX column as described in Example 1(b). The protein and IgG yields of the differently loaded solutions were compared. The results are shown in figure 1 middle. Comparing the effect on IgG yield of 1M Tris buffer at different pH switches with 0.2M NaOH used in the prior art method de...
Embodiment 3
[0096] This example shows the effect of the concentration of Tris buffer used on the desired pH transition to 5.8. Starting from a solution incubated at low pH 4, the pH was shifted from 4 to 5.8 using 1M, 0.5M and 0.25M Tris buffer, respectively.
[0097] The results are shown in figure 2 middle. Independently of the starting material (sourced PPT-II+III, NA or recovered NA), IgG loss decreased with increasing molarity of Tris buffer used.
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