Protein Extraction and Immunoblotting Protocol

Preparation of cell lysates, extraction of proteins, and immunoblotting were carried out as described earlier (32 (link), 34 (link)). Proteins were separated on TRIS-glycine gels (7.5% for NOS2 and 12% for Arginase 1) and transferred to PVDF membranes. Staining with appropriate primary and secondary antibodies was followed by signal visualization using the Chemo Star Imager (Intas Science Imaging Instruments, Göttingen, Germany). Densitometry of signals was done using ImageJ (Version 1.52a; Rasband, W., ImageJ, National Institutes of Health, USA, https://imagej.nih.gov/ij/).

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Frick L., Hinterland L., Renner K., Vogl M., Babl N., Heckscher S., Weigert A., Weiß S., Gläsner J., Berger R., Oefner P.J., Dettmer K., Kreutz M., Schatz V, & Jantsch J. (2022). Acidic Microenvironments Found in Cutaneous Leishmania Lesions Curtail NO-Dependent Antiparasitic Macrophage Activity. Frontiers in Immunology, 13, 789366.

Publication 2022

Chemo Star Imager

Antibodies Arginase 1 Cell Densitometry Gels Glycine Membranes Nos2 Proteins Pvdf Tris

Corresponding Organization : University of Regensburg

Top 5 similar protocols

Variable analysis

independent variables

Preparation of cell lysates
Extraction of proteins
Immunoblotting

dependent variables

Protein expression levels of NOS2 and Arginase 1

control variables

Protein separation on TRIS-glycine gels (7.5% for NOS2 and 12% for Arginase 1)
Transfer of proteins to PVDF membranes
Staining with appropriate primary and secondary antibodies
Signal visualization using the Chemo Star Imager
Densitometry of signals using ImageJ

controls

Positive and negative controls not explicitly mentioned

Annotations

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