Gfp tag

Manufactured by Proteintech

Sourced in United States

The GFP-tag is a protein tag used for fluorescent labeling of proteins. It is derived from the green fluorescent protein (GFP) found in the jellyfish Aequorea victoria. The GFP-tag can be fused to the N-terminus or C-terminus of a target protein, allowing the visualization and tracking of the tagged protein within cells or tissues.

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Lab products found in correlation

β actin, by Proteintech (5 mentions) Gapdh, by Proteintech (3 mentions) Rc dc assay, by Bio-Rad (2 mentions) α tubulin, by Proteintech (2 mentions) Myc tag, by Proteintech (2 mentions) Flag tag (flag), by Proteintech (2 mentions) Gw4869, by Merck Group (1 mentions) P44 42 mapk erk1 2 l34f12, by Cell Signaling Technology (1 mentions) P akt ser473 d9e xp, by Cell Signaling Technology (1 mentions) Alexa fluor 555 goat anti mouse igg, by Thermo Fisher Scientific (1 mentions) Flag m2, by Merck Group (1 mentions) Goat anti rabbit igg secondary antibody, by Boster Bio (1 mentions) Bcl 2, by Abmart (1 mentions) Alpha tubulin, by Abcam (1 mentions) Keap1, by Proteintech (1 mentions) Immpresstmhrp anti rabbit igg, by Vector Laboratories (1 mentions) Smurf1, by Abcam (1 mentions) Smurf1, by Santa Cruz Biotechnology (1 mentions) Eif2α, by ABclonal (1 mentions) Histone h2b, by Santa Cruz Biotechnology (1 mentions)

8 protocols using gfp tag

Immunoblotting Analysis of ER Stress Markers

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Mammalian cells were lysed in radioimmunoprecipitation assay buffer (RIPA) buffer with the addition of protease (Roche cOmplete), and phosphatase inhibitors (GB Sciences). Protein levels of lysates were determined using the Bio-Rad DC/RC assay. Equal amounts of protein lysate were boiled with SDS load buffer, and equal amounts of protein were loaded. Immunoblotting was performed with the following antibodies: GAPDH (Cat, no. 60004-1-lg; Proteintech), ATF6 rabbit polyclonal (Cat. no. 24169-1-AP; Proteintech), ATF6 mouse monoclonal (Cat. no. 66563-1-Ig; Proteintech), β-Actin (Cat. no. 20536-1-AP; Proteintech), α-Tubulin (Cat. no. 66031-1-Ig; Proteintech), SEL1L (Cat. no. ab78298; Abcam), ABCD3 (Cat. no. 66697-1-Ig; Proteintech), GFP tag (Cat. no. 66002-1-Ig; Proteintech), BiP (Cat. no. 11587-1-AP; Proteintech), ATF4 (Cat. no. 11815S; Cell Signaling Technology).

Ibe N.U., Subramanian A, & Mukherjee S. (2021). Non-canonical activation of the ER stress sensor ATF6 by Legionella pneumophila effectors. Life Science Alliance, 4(12), e202101247.

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Measuring Ras Signaling Pathway

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The following antibodies used to measure Ras signaling were purchased from Cell Signaling Technology (Danvers, MA): pAkt (Ser473) (D9E) XP (Cat#4060), total Akt (pan) (40D4) (Cat#2920), p-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP (Cat#4370) and p44/42 MAPK (Erk1/2) (L34F12) (Cat#4696). The following antibodies used to measure housekeeping genes were purchased from Proteintech Group (Rosemont, IL): β-actin (Cat#60008-1-1 g) and GFP-tag (Cat#66002-1-lg; clone #1E10H7). cDNA of human SMPD1 (GenScript, Cat# OHu18710D), SMPD2 (GenScript, Cat# OHu30447D) and SMPD3 (Sino Biological, Cat# HG15755-G) were cloned into GFP-N1 vector. CellLight Lysosomes-RFP (Cat#C10597), Early endosomes-RFP (Cat#C10587), Late Endosomes-RFP (Cat#C10589), Golgi-RFP (Cat#10593), Mitochondria-RFP (Cat#C10601) and ER-tracker (E34250) were purchased from Invitrogen. Altenusin (Cat#SML2193) and GW4869 (Cat#567715) were purchased from MilliporeSigma. Avicin compounds were provided in collaboration with Dr. Jordan Gutterman (MD Anderson Cancer Center; Houston, TX).

Garrido C.M., Henkels K.M., Rehl K.M., Liang H., Zhou Y., Gutterman J.U, & Cho K.J. (2020). Avicin G is a potent sphingomyelinase inhibitor and blocks oncogenic K- and H-Ras signaling. Scientific Reports, 10, 9120.

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Comprehensive Immunoblotting Antibody Panel

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Primary antibodies used against proteins were as follow: SMURF1 (Santa Cruz, sc-100616), SMURF1 (Abcam, ab57573), BiP/GRP78 (ABclonal, A0241); Actin (Sigma, A1978), p62 (MBL, PM045); LC3B (Sigma, L7543); Phospho-IRE1(S724) (ABclonal, AP0878); IRE1(ABclonal, A17940); Phospho-JNK1/2/3 (T183+T183+T221) (Abmart, T55541); JNK (ABclonal, A4867); Phospho-eIF2α (Ser51) (ABclonal, AP0692); eIF2α (ABclonal, A0764); XBP1 (ABclonal, A1731); ATF4 (Santa Cruz, sc-390063); CHOP (ABclonal, A6504); BCL-2 (ABmart, T40056); Flag M2 (Sigma, F3165); GFP-tag (Proteintech, 66002-1-lg); HA-tag (MBL, M180-3); Myc-Tag (Proteintech, 16286-1-AP); NRF2 (Proteintech, 16396-1-AP); KEAP1 (Proteintech, 10503-2-AP); Ubiquitin (MBL, D058-3); alpha Tubulin (Abcam, ab7291), Histone H2B (Santa Cruz, sc-515808); Caspase3 (Santa Cruz, sc-7272); Ki67 (Abcam, ab16667).
Secondary antibodies used were as follow: goat anti-mouse IgG secondary antibody (BOSTER, BA1050); goat anti-rabbit IgG secondary antibody (BOSTER, BA1054); Alexa Fluor^® 555 goat anti-mouse IgG (Life Technologies, A21425); ImmPRESS^TM HRP anti-Rabbit IgG (VECTOR, MP-7401); ImmPRESS^TM HRP anti-Mouse IgG (VECTOR, MP-7402); Rabbit anti-mouse IgG (CST, 58802) and Mouse anti-Rabbit IgG (CST, 93702) were used to avoid interference of the IgG heavy chain.

Dong L., Xu M., Li Y., Xu W., Wu C., Zheng H., Xiao Z., Sun G., Ding L., Li X., Li W., Zhou L, & Xia Q. (2023). SMURF1 attenuates endoplasmic reticulum stress by promoting the degradation of KEAP1 to activate NRF2 antioxidant pathway. Cell Death & Disease, 14(6), 361.

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Western Blotting Technique for Protein Detection

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First, we used Radioimmunoprecipitation assay (RIPA) lysis buffer (Beyotime, Jiangsu, China) to lyse cells to obtain proteins, which were quantified using the Bradford method. The proteins were separated using SDS-polyacrylamide gel electrophoresis (SDS-PAGE, Beyotime) and transferred to polyvinylidene fluoride (PVDF) membranes (Millipore, Billerica, MA, USA). After blocking the membranes for 1 h at room temperature with 5% skim milk, the membranes were incubated with the corresponding primary antibodies at 4 °C overnight. The primary antibodies recognized MFSD4A (Genetex, Irvine, CA, USA), EPHA2 (CST, Danvers, MA, USA), green fluorescent protein (GFP)-tag (Proteintech, USA), HA-tag (Proteintech, Rosemont, IL, USA), Flag-tag (Proteintech), Vimentin (CST), E-cadherin (CST), N-cadherin (CST), Slug (CST), Snail (CST), AKT (CST), phosphorylated (p)-AKT (CST), ERK1/2 (Abcam, Cambridge, MA, USA), p-ERK1/2 (CST), PI3K (Abcam), p-PI3K (CST) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (Proteintech). The next day, the corresponding secondary antibodies were added and incubated with the membranes for 1 h at room temperature. Finally, enhanced chemiluminescence was used to observe the results.

Yang H., Qin G., Luo Z., Kong X., Gan C., Zhang R, & Jiang W. (2022). MFSD4A inhibits the malignant progression of nasopharyngeal carcinoma by targeting EPHA2. Cell Death & Disease, 13(4), 332.

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Measuring Ras Signaling with Validated Antibodies

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The following antibodies to measure Ras signaling were purchased from Cell Signaling Technology: pAkt (Ser473) (Cat# 4060), total Akt (Cat# 2920), ppERK (Thr202/Tyr204) (Cat# 4370), total ERK (Cat# 4696), p-p90RSK (Cat #9344), and total RSK (Cat #9355). The following antibodies used to measure housekeeping genes were purchased from Proteintech: β-actin (Cat# 66009-1-Ig), GFP-tag (Cat# 60002-1-Ig), α-tubulin (Cat# 11224-1-AP), BACH1 (Cat# 66762-1-IG), H-Ras (Cat# 18295-1-AP), K-Ras2B (Cat# 16155-1-AP), and K-Ras2A (Cat #16156-1). N-Ras antibody (F155) (Cat #sc-31) was purchased from Santa Cruz Biotechnology. The RFP-tagged organelle markers were purchased from Invitrogen: CellLight Golgi-RFP BacMam 2.0 (Cat# C10593), CellLight Lysosomes-RFP BacMam 2.0 (Cat #C10597), CellLight Mitochondria-RFP BacMam 2.0 (Cat #C10601), CellLight Late Endosome-RFP BacMam 2.0 (Cat #C10589), CellLight Early Endosome-RFP BacMam 2.0 (Cat #C10587), and ER-Tracker Red (Cat #E34250; BODIPY TR Glibenclamide). Mitomycin C was purchased from Alfa Aesar Co. (Cat #J63193). Carmustine was purchased from MedChemExpres (Cat # HY-13585/CS-2935). N-acetylcysteine was purchased from Sigma-Aldrich (Cat # A7250-10G). Hydrogen peroxide (H₂O₂) was purchased from Sigma-Aldrich (Cat #H1009).

Rehl K.M., Selvakumar J., Pitsch R.L., Hoang D., Arumugam K., Harshman S.W., Gorfe A.A, & Cho K.J. (2023). A new ferrocene derivative blocks K-Ras localization and function by oxidative modification at His95. Life Science Alliance, 6(11), e202302094.

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Immunoprecipitation and Western Blotting Protocol

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For western blotting, the detailed protocol was previously described.⁶³ (link) For IP, podocytes were lysed in IP lysis buffer before being briefly sonicated, and the supernatants were incubated with antibodies and protein A/G-Sepharose beads after centrifugation. Finally, the immunocomplexes were washed and immunoblotted with antibodies. The following antibodies were used: ZO-1 (Invitrogen, Calrsbad, CA, USA), Desmin, podocin, Sox4, p21^cip1/waf1 (Abcam, Cambridge, MA, USA), β-actin, synaptopodin, cdc2, CyclinB, p53, GFP-tag, hemagglutinin (HA)-tag, Myc-tag, FLAG tag, and HRP-conjugated goat anti-rabbit/mouse IgG (Proteintech, Rosemont, IL, USA).

Wang Z., Chang Y., Liu Y., Liu B., Zhen J., Li X., Lin J., Yu Q., Lv Z, & Wang R. (2022). Inhibition of the lncRNA MIAT prevents podocyte injury and mitotic catastrophe in diabetic nephropathy. Molecular Therapy. Nucleic Acids, 28, 136-153.

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Immunoblotting of Cellular Proteins

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Total protein was isolated from cells using lysis buffer (Beyotime, China). Immunoblotting was performed with primary antibodies against FBXW7 (Proteintech, China), MAP4 (Proteintech, China), MMP3 (Proteintech, China), VEGFA (Proteintech, China), ERK (CST, USA), phosphorylated ERK (Thr202/Tyr204) (CST, USA), GAPDH (Proteintech, China), His-tag (Proteintech, China), and GFP-tag (Proteintech, China). Secondary antibodies were purchased from CWbiotech (China). The signal was visualized using super enhanced chemiluminescence (ECL) detection reagent (Tanon, China).

Pan Y., Liu J., Gao Y., Guo Y., Wang C., Liang Z., Wu M., Qian Y., Li Y., Shen J., Lu C, & Ma S. (2023). FBXW7 loss of function promotes esophageal squamous cell carcinoma progression via elevating MAP4 and ERK phosphorylation. Journal of Experimental & Clinical Cancer Research : CR, 42, 75.

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Immunoblotting Assay for Cellular Stress Markers

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Mammalian cells were lysed in Radioimmunoprecipitation assay buffer (RIPA) buffer with the addition of protease (Roche cOmplete), and phosphatase inhibitors (GB Sciences). Protein levels of lysates were determined using the BioRad DC/RC assay. Equal amounts of protein lysate were boiled with SDS load buffer, and equal amounts of protein were loaded. Immunoblotting was performed with the following antibodies: GAPDH (Proteintech, Cat: 60004-1-lg), ATF6 (Proteintech, Cat: 60004-1-lg), beta-Actin (Proteintech, Cat: 20536-1-AP), alpha-Tubulin (Proteintech, Cat: 66031-1-Ig), Sel1L (Abcam, Cat: ab78298), GFP tag (Proteintech, Cat: 66002-1-Ig), BiP (Proteintech, Cat: 11587-1-AP), ATF4 (Cell Signaling Technology, Cat: 11815S) .

Ibe N.U., & Mukherjee S. (2020). Non-canonical activation of the ER stress sensor ATF6 byLegionella pneumophilaeffectors.

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