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M1 monoclonal flag antibody

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The M1 monoclonal FLAG antibody is a laboratory reagent used for the detection and purification of proteins tagged with the FLAG epitope. The antibody is highly specific and binds to the FLAG peptide sequence with high affinity, allowing for the identification and isolation of FLAG-tagged proteins from complex mixtures.

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Lab products found in correlation

Du897 emccd camera, by Oxford Instruments (4 mentions) Alexa 647 dye, by Thermo Fisher Scientific (4 mentions) Ti e inverted microscope, by Nikon (4 mentions) 35 mm glass bottomed culture dishes, by MatTek (4 mentions) Poly l lysine (pll), by Merck Group (4 mentions) Nis elements software, by Nikon (2 mentions)

4 protocols using m1 monoclonal flag antibody

1

Super-resolution Imaging of Cellular Proteins

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PALM was performed at 37°C using a Nikon Ti-E inverted microscope equipped for through-the-objective TIR-FM and outfitted with a temperature-, humidity-, and CO2-controlled chamber (Okolab). Images were obtained with an Apo TIRF 100X, 1.49 numerical aperture objective (Nikon) with solid-state lasers of 405, 488, 561, and 647 nm (Keysight Technologies) as light sources. An Andor iXon DU897 EMCCD camera controlled by NIS-Elements 4.1 software was used to acquire 5000 periods with continuous activation by 405 nm light. Cells were transfected as indicated according to manufacture protocol 48 hours before imaging and then plated on poly-L-lysine (0.0001%, Sigma) coated 35-mm glass-bottomed culture dishes (MatTek Corporation) 24 hours before imaging. Cells were labeled with M1 monoclonal FLAG antibody (1:1000, Sigma F-3040) conjugated to Alexa 647 dye (Life Technologies) for 10 minutes at 37°C, washed one time with imaging media (20 mM HEPES, 135 mM NaCl, 5mM D+ glucose, 5 mM KCl, 0.4 mM MgCl2, 1.8 mM CaCl2, pH 7.4), and imaged live in imaging media. NIS Elements software (Nikon) was used to resolve and analyze the images. Full width at half max calculations were used to measure size of the super resolved structures. PALM imaging experiments were performed at least three independent times.
Eichel K., Jullié D, & von Zastrow M. (2016). β-arrestin drives MAP kinase signaling from clathrin-coated structures after GPCR dissociation. Nature cell biology, 18(3), 303-310.
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2

Live-cell TIRF Microscopy of HEK293 Cells

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TIRF microscopy was performed at 37°C using a Nikon Ti-E inverted microscope equipped for through-the-objective TIRF microscopy and outfitted with a temperature-, humidity-, and CO2-controlled chamber (Okolab). Images were obtained with an Apo TIRF 100X, 1.49 numerical aperture objective (Nikon) with solid-state lasers of 405, 488, 561, and 647 nm (Keysight Technologies). An Andor iXon DU897 EMCCD camera controlled by NIS-Elements 4.1 software was used to acquire image sequences every 2 seconds for 10 minutes. Unless indicated otherwise, live cell microscopy assays were performed using HEK 293 cells. Cells were transfected as indicated according to manufacturer’s protocol 48 hours before imaging and then plated on poly-L-lysine (0.0001%, Sigma) coated 35-mm glass-bottomed culture dishes (MatTek Corporation) 24 hours before imaging. Cells were labeled with M1 monoclonal FLAG antibody (1:1000, Sigma F-3040) conjugated to Alexa 647 dye (Life Technologies) for 10 minutes at 37°C prior to imaging, washed, and imaged live in DMEM without phenol red (UCSF Cell Culture Facility) supplemented with 30 mM HEPES, pH 7.4 (UCSF Cell Culture Facility). Cells were treated with bath application of 10 μM of the indicated agonist at time equals 0 seconds for experiments shown as timecourses. At least three independent experiments were performed for all live cell TIRF microscopy imaging.
Eichel K., Jullié D., Barsi-Rhyne B., Latorraca N.R., Masureel M., Sibarita J.B., Dror R.O, & von Zastrow M. (2018). Catalytic activation of β-arrestin by GPCRs. Nature, 557(7705), 381-386.
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3

Live-cell TIRF Microscopy of HEK293 Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols
TIRF microscopy was performed at 37°C using a Nikon Ti-E inverted microscope equipped for through-the-objective TIRF microscopy and outfitted with a temperature-, humidity-, and CO2-controlled chamber (Okolab). Images were obtained with an Apo TIRF 100X, 1.49 numerical aperture objective (Nikon) with solid-state lasers of 405, 488, 561, and 647 nm (Keysight Technologies). An Andor iXon DU897 EMCCD camera controlled by NIS-Elements 4.1 software was used to acquire image sequences every 2 seconds for 10 minutes. Unless indicated otherwise, live cell microscopy assays were performed using HEK 293 cells. Cells were transfected as indicated according to manufacturer’s protocol 48 hours before imaging and then plated on poly-L-lysine (0.0001%, Sigma) coated 35-mm glass-bottomed culture dishes (MatTek Corporation) 24 hours before imaging. Cells were labeled with M1 monoclonal FLAG antibody (1:1000, Sigma F-3040) conjugated to Alexa 647 dye (Life Technologies) for 10 minutes at 37°C prior to imaging, washed, and imaged live in DMEM without phenol red (UCSF Cell Culture Facility) supplemented with 30 mM HEPES, pH 7.4 (UCSF Cell Culture Facility). Cells were treated with bath application of 10 μM of the indicated agonist at time equals 0 seconds for experiments shown as timecourses. At least three independent experiments were performed for all live cell TIRF microscopy imaging.
Eichel K., Jullié D., Barsi-Rhyne B., Latorraca N.R., Masureel M., Sibarita J.B., Dror R.O, & von Zastrow M. (2018). Catalytic activation of β-arrestin by GPCRs. Nature, 557(7705), 381-386.
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4

Super-resolution Imaging of Cellular Proteins

Check if the same lab product or an alternative is used in the 5 most similar protocols
PALM was performed at 37°C using a Nikon Ti-E inverted microscope equipped for through-the-objective TIR-FM and outfitted with a temperature-, humidity-, and CO2-controlled chamber (Okolab). Images were obtained with an Apo TIRF 100X, 1.49 numerical aperture objective (Nikon) with solid-state lasers of 405, 488, 561, and 647 nm (Keysight Technologies) as light sources. An Andor iXon DU897 EMCCD camera controlled by NIS-Elements 4.1 software was used to acquire 5000 periods with continuous activation by 405 nm light. Cells were transfected as indicated according to manufacture protocol 48 hours before imaging and then plated on poly-L-lysine (0.0001%, Sigma) coated 35-mm glass-bottomed culture dishes (MatTek Corporation) 24 hours before imaging. Cells were labeled with M1 monoclonal FLAG antibody (1:1000, Sigma F-3040) conjugated to Alexa 647 dye (Life Technologies) for 10 minutes at 37°C, washed one time with imaging media (20 mM HEPES, 135 mM NaCl, 5mM D+ glucose, 5 mM KCl, 0.4 mM MgCl2, 1.8 mM CaCl2, pH 7.4), and imaged live in imaging media. NIS Elements software (Nikon) was used to resolve and analyze the images. Full width at half max calculations were used to measure size of the super resolved structures. PALM imaging experiments were performed at least three independent times.
Eichel K., Jullié D, & von Zastrow M. (2016). β-arrestin drives MAP kinase signaling from clathrin-coated structures after GPCR dissociation. Nature cell biology, 18(3), 303-310.
+ Open protocol
+ Expand

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