Nanoparticles in Biology and Medicine by Unknown

Nanoparticles in Biology and Medicine by Unknown

Author:Unknown
Language: eng
Format: epub
ISBN: 9781071603192
Publisher: Springer US


11.Al protein preparations will benefit from buffer exchange. “Difficult” proteins will certainly need a buffer exchange to 0.01–0.1% sodium citrate to help stabilize AuNPs.

12.To scale up purification use two spin column. 100 μl of GST fusion protein at a concentration of ~0.1 mg/ml will be sufficient to saturate at least 1 ml of 1 O.D. AuNPs with protein and will likely provide an excess of protein [15].

13.Immobilization of proteins may affect AuNP colloidal stability . To ensure that all AuNP surface is covered with proteins and to improve AuNP stability during centrifugal washing steps and downstream processing, AuNPs should be blocked with excess of irrelevant protein. This applies to AuNP–protein conjugates as well as to AuNPs without any immobilized protein. Since the experiment described here relies on the measurement of the functional activity of Staphylokinase, the presence of BSA is not expected to affect the readout; hence, BSA could be used for blocking AuNPs. A control sample AuNP-BSA has also been generated and used in the downstream functional testing to account for any effects due to the presence of BSA alone. The choice of blocking method will depend on the proteins immobilized and the property being measured. A short incubation time may be added, albeit the excess of protein used (100 μl of 1% BSA per 1 ml of AuNP), no additional incubation time was necessary, prior to the AuNP washing step.



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