Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2021 Oct 26;11:21048. doi: 10.1038/s41598-021-99887-5

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© The Author(s) 2021

Open Access This 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/.

PMC Copyright notice

Immunoblotting studies on boid cells transfected with different arenaviral NPs. (a–d) Immunoblotting analyses of whole-cell lysates (Tot) and mitochondrial preparations (Mit) obtained from Boa constrictor V/4Br cells transfected with constructs expressing wt or mutUGV1-NP-FLAG (a), UHV1-NP-FLAG (b), wt or mutJUNV-NP-HA, or LCMV-NP-HA (c) and HISV1-NP-FLAG (d), all at three dpt. Non-transfected (Mock) samples were used as negative controls. About 1/5 of the transfected cells were used for whole-cell lysate procedure (Tot) and the other 4/5 for mitochondria isolation (Mit). 20 µg (a,b) or 12 µg (c,d) of protein per sample from both Tot and Mit were loaded on standard SDS-PAGE gels followed by immunoblotting analyses. The nitrocellulose membranes were incubated sequentially with the following antibodies in the presented order: (1) mouse anti-FLAG tag 1:500 (a,b,d) or mouse anti-HA tag 1:500 (c); (2) rabbit anti-MTS-NP 1:200 (a–c), or rabbit anti-Hartmani-NP 1:500 (d); (3) mouse anti-tubulin 1:500 (a–d); (4) mouse anti-MFN2 1:200 (a,b,d) or 1:100 (c). Anti-tubulin and anti-MFN2 specific signals at known molecular weight did not require membrane stripping, except for panel c, where a stripping step was introduced before probing with mouse anti-MFN2. Tubulin and MFN2 (both in red, secondary antibody: IRDye 680RD Donkey anti-mouse) were used as cytosolic and mitochondrial marker, respectively. Arenavirus NPs (63–68 kDa) are indicated (black arrows). (a,b) Left panels: FLAG tag in red (secondary antibody: IRDye 680RD Donkey anti-mouse); middle panels: MTS-NP in green (secondary antibody: IRDye 800CW Donkey anti-rabbit); right panels: merged image. (c) Left panel: HA tag in red (secondary antibody: IRDye 680RD Donkey anti-mouse); middle panel: MTS-NP in green (secondary antibody: IRDye 800CW Donkey anti-rabbit); right panel: merged image. (d) Left panel: FLAG tag in red (secondary antibody: IRDye 680RD Donkey anti-mouse); middle panel: Hartmani-NP in green (secondary antibody: IRDye 800CW Donkey anti-rabbit); right panel: merged image. Immunodetection was performed using the Odyssey Infrared Imaging System (LICOR, Biosciences) providing also the molecular marker (Precision Plus Protein Dual Color Standards, Bio-Rad) used. Full-length blots are presented in Supplementary Fig. S3.