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We thank Dr. Yael Yoffe for providing human embryonic stem cell extracts and Prof. Orly Reiner for the mouse protein extracts.
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In sham-operated mice, protein extracts were prepared from equivalent brain regions.
For healthy mice, protein extracts from human testis were used as positive controls (Applied Biosystems, Foster City, CA, USA) [ 37].
The light and heavy dimethyl-labeled peptides derived from equal amount of wild-type and HDAC6 knockout mouse liver protein extracts were pooled and desalted using C18 microcolumns.
To further clarify these binding relationships in vivo, chromatographic fractionation of mouse testis protein extracts was performed to purify the putative complex es) containing PPP1R11, PP1γ2 and Sds22.
HeLa cell or mouse liver protein extracts were analyzed by Western blotting.
Alexa 488 Fluorescent Hydroxylamine (FHA, Invitrogen) was added to mouse brain protein extracts at 1 mg of FHA for 20 mg of protein.
7 Mouse brain protein extract was systematically included to ensure Nogo-A band identification.
Twenty micrograms of total cell lysate (CHO or HEK293) or mouse chondrocyte protein extract were separated by SDS PAGE on 4 12% Bis-Tris gels (Invitrogen).
In addition, we report the proteomics analyses of complex biological samples using RP-HILIC with mouse chondrogenic cell protein extracts and deglycosylated human serum.
In fact, it has been shown that mouse protein is easily extracted from brain by high salt buffer, instead human parkin is only extracted with harsher buffers, especially in elderly.
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