Improve Research Reproducibility A Bio-protocol resource
Concise Method
tooltip

Western blot

AP Arie Passov
MI Minna Ilmakunnas
MP Marjut Pihlajoki
KH Kethe Hermunen
ML Marko Lempinen
IH Ilkka Helanterä
VK Villemikko Kailari
MH Markku Heikinheimo
SA Sture Andersson
EP Eero Pesonen
ask Ask a question
Favorite

Non-reducing Western blotting (n = 25) was performed on all available samples, to detect plasma NGAL isoforms in preoperative blood sample and in renal venous blood at reperfusion. The method has been described in detail previously [22]. In brief, the samples were mixed 1:4 with Laemmli sample buffer. Protein content was quantified and 10 µg of protein was separated by electrophoresis using Mini-Protean TGX Stain-Free gels (Bio-Rad Laboratories, CA, USA) and transferred onto a polyvinylidene fluoride membrane (Thermo Fisher Scientific, MA, USA). Primary antibody used was a polyclonal rabbit anti-HNL-NGAL (Diagnostics Development, Uppsala, Sweden) at dilution of 1:1000. Normalization and quantification of the protein band intensity were carried out using Image Lab 6.0 software.

Copyright and License information: The Author(s) ©2021
Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

Do you have any questions about this protocol?

Post your question to gather feedback from the community. We will also invite the authors of this article to respond.

post Post a Question
0 Q&A