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Immunoblotting

CK Christine Koch
YK Younghoon Kim
TZ Tobias Zöller
CB Christina Born
AS Alexander Steinle
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For immunoblot analysis, purified NK cells from spleens of naïve mice were lysed using Pierce RIPA buffer (Thermo Fisher Scientific), containing the Complete protease inhibitor cocktail (Roche, Mannheim, Germany). 20 µg of total lysates were separated via non-reducing SDS-PAGE and transferred to PVDF membranes (Carl Roth, Arlesheim, Switzerland) by semi-dry blotting. Membranes were probed with 0.5 µg/ml anti-CD3ζ (6B10.2) at 4°C for 16 h, followed by detection with horseradish peroxidase-conjugated goat anti-mouse IgG antibody (Jackson ImmunoResearch Laboratories, West Grove, PA, USA). Immunoblot signals were generated with Femto-ECL© (Thermo Fisher Scientific).

Copyright and License information:  ©2017 Koch, Kim, Zöller, Born and Steinle.
This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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