STED Microscopy Immunostaining Protocol

Fixation and staining of the cells for stimulated emission depletion (STED) microscopy have been described previously [23 (link)]. Incubation with the primary antibody was performed overnight at 4 °C. The following antibodies were used: anti-Ku70 (#ab 83501) (Abcam, Berlin, Germany) and anti-IKKγ/NEMO (#SAB1404591) (Sigma-Aldrich, Taufkirchen, Germany). The following secondary antibodies were used: anti-rabbit ATTO 647N (dilution 1:200) and anti-mouse Chromeo 494 (dilution 1:50) (Active Motif, La Hulpe, Belgium).

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Medunjanin S., Putzier M., Nöthen T., Weinert S., Kähne T., Luani B., Zuschratter W, & Braun-Dullaeus R.C. (2020). DNA-PK: gatekeeper for IKKγ/NEMO nucleocytoplasmic shuttling in genotoxic stress-induced NF-kappaB activation. Cellular and Molecular Life Sciences, 77(20), 4133-4142.

Publication 2020

anti-IKKγ/NEMO ATTO 647N Chromeo 494

Antibodies Antibody Cells Dilution Ikkγ Ku70 Microscopy Mouse Rabbit

Corresponding Organization : University Hospital Magdeburg

Other organizations : Leibniz Institute for Neurobiology

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Variable analysis

independent variables

Primary antibodies: anti-Ku70 (#ab 83501) (Abcam, Berlin, Germany) and anti-IKKγ/NEMO (#SAB1404591) (Sigma-Aldrich, Taufkirchen, Germany)

dependent variables

Not explicitly mentioned

control variables

Fixation and staining of the cells for stimulated emission depletion (STED) microscopy
Incubation with the primary antibody performed overnight at 4 °C
Secondary antibodies: anti-rabbit ATTO 647N (dilution 1:200) and anti-mouse Chromeo 494 (dilution 1:50) (Active Motif, La Hulpe, Belgium)

controls

Positive control: Not mentioned
Negative control: Not mentioned

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