α-CD8 and rabbit α-HA were obtained from Sigma, α-BAP31, α-Actin and α-tubulin from AbCam, α-ERGIC53 from Alexis, mouse α-HA from Santa Cruz, α-EEA1 from Cell signalling and α-GM130 from BD biosciences. IRDye 800 CW and IRDye 680 RD antibodies were from LI-COR, and fluorescently conjugated secondary antibodies for microscopy were from Jackson Laboratories (Stratech Scientific). The inhibitors leupeptin (Enzo Life Sciences), pepstatin A (Sigma), Z-LLF-CHO (PSII, Calbiochem), and cycloheximide (CHX, Sigma) were used at a final concentration of 0.5 mM, 1 μg/ml, 10 μM, and 100 μg/ml respectively.
α gm130
Manufactured by BD
Sourced in United States
α-GM130 is a lab equipment product used to detect the GM130 protein, a Golgi matrix protein involved in vesicle tethering and Golgi apparatus structure. The product is designed to be used in research applications.
Automatically generated - may contain errors
Lab products found in correlation
710 confocal microscope, by Zeiss (3 mentions)
Leupeptin, by Enzo Life Sciences (1 mentions)
α tubulin, by Abcam (1 mentions)
α actin, by Abcam (1 mentions)
Mouse α ha, by Santa Cruz Biotechnology (1 mentions)
Pepstatin a, by Merck Group (1 mentions)
Irdye 800cw, by LI COR (1 mentions)
Irdye 680rd, by LI COR (1 mentions)
Cycloheximide (chx), by Merck Group (1 mentions)
α myc tag, by Merck Group (1 mentions)
α myc tag, by Cell Signaling Technology (1 mentions)
α eea1, by Abcam (1 mentions)
Fv1000 confocal microscope, by Zeiss (1 mentions)
α lc3b, by Cell Signaling Technology (1 mentions)
α β actin, by Merck Group (1 mentions)
α β catenin, by Cell Signaling Technology (1 mentions)
αγ tubulin, by Merck Group (1 mentions)
3 protocols using α gm130
1
Antibody Sources and Inhibitor Use
2
Immunofluorescence Staining and Live-Cell Imaging
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For Immunofluorescence studies, cells were grown on glass coverslips for 2 days, then the coverslips were washed once in PBS and then fixed with 4% p-formaldehyde for 30 min at room temperature. Immunofluorescence staining of cells was carried out as previously described.39 (link) An Olympus FV1000 confocal microscope with a 60 × objective and a Zeiss 710 confocal microscope with × 63 objective were used for taking images.
Antibodies used were as follows: α-Myc-Tag (Millipore; 4A6), α-Myc-Tag (Cell Signaling; #2272), α-LC3B (Cell Signaling; #2775), α-LAMP2 (BD Pharmingen, San Jose, CA, USA; CD107b), α-EEA1 (Abcam, ab2900), α-Calnexin (Cell Signaling; #2679), α-WIPI2 (Bio-rad; MCA578OGA) and α-GM130 (BD Bioscience, #610822).
For live-cell imaging, cells were transfected with GFP-tagged DRAM-3 and RFP-tagged Rab5 or Rab7 at least 2 days before confocal analysis. An Andor spinning disc confocal system with Cairn Optsplit (for simultaneous GFP/RFP imaging) was used for imaging. Videos of endosomes were taken for 1 min, with pictures in 1-s intervals.
Antibodies used were as follows: α-Myc-Tag (Millipore; 4A6), α-Myc-Tag (Cell Signaling; #2272), α-LC3B (Cell Signaling; #2775), α-LAMP2 (BD Pharmingen, San Jose, CA, USA; CD107b), α-EEA1 (Abcam, ab2900), α-Calnexin (Cell Signaling; #2679), α-WIPI2 (Bio-rad; MCA578OGA) and α-GM130 (BD Bioscience, #610822).
For live-cell imaging, cells were transfected with GFP-tagged DRAM-3 and RFP-tagged Rab5 or Rab7 at least 2 days before confocal analysis. An Andor spinning disc confocal system with Cairn Optsplit (for simultaneous GFP/RFP imaging) was used for imaging. Videos of endosomes were taken for 1 min, with pictures in 1-s intervals.
3
Immunofluorescence Centrosome Markers Protocol
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Primary antibodies used for immunofluorescent markers of the centrosome were α-GM130 (BD 610822, 1:1,000 dilution [dil.]) and α-γ-tubulin (Sigma Aldrich T5326-25UL, 1:1,000). The secondary used for both stains was α-Ms-Alexa-488 (Invitrogen A28175, 1:1,000). Tissue fixation and staining was carried out using standard protocols using cold methanol (52 ). Immunofluorescent samples were imaged using confocal microscopy (see below). Antibodies used for Western blotting and immunofluorescence were α-β-catenin (Cell Signaling, #2698S, 1:1,000) and α-β-actin (Sigma, A3853, 1:1,000). Secondary antibodies used were α-Gt-680RD and α-Ms-800CW (Licor 926-6807 and 926-32212, respectively, both 1:10,000 dil.). Standard immunoblot procedures were used (53 ).
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