NIH3T3 (ATCC CRL-1658) or HEK-293 cells (ATCC CRL-1573) were maintained in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 10% (v/v) fetal bovine serum (FBS), 1000 U/ml penicillin, 1000 μg/ml streptomycin, and 20 mM L-Glutamine (all Gibco) at 37°C and 5% CO2. NIH3T3 cells were grown on coverslips in 6-well plates and transfected using EndoFectinTM Max Transfection Reagent, following the recommendations of the manufacturer (GeneCopoeia). Twenty-four hours after transfection, cells were washed in phosphate-buffered saline (PBS) and fixed in methanol for 10 min at −20°C. Specimens were then permeabilized in 0.3% TritonX-100 in PBS for 10 min at room temperature and blocked for 1 h using blocking solution (PBS containing 1% BSA and 0.3% TritonX-100). Samples were incubated with primary antibodies toward CCDC42 (ARP52735_P050, antibodies-online ABIN2785068), Pericentrin (PRB432C, Covance), acetylated tubulin (6-11B-1; Sigma-Aldrich), gamma-tubulin (GTU-88, Sigma-Aldrich), ODF1 (ABIN4341345, antibodies-online), and GFP (raised in rabbit, self-made) at 37°C for 1 h. Secondary antibodies used are goat anti-mouse-IgG DyLight 488 (#35503, Thermo Fisher Scientific), and goat anti-rabbit MFP590 (#MFP-A1037, Mobitec). DNA was counterstained with DAPI. Images were taken by confocal microscopy (LSM 780, Zeiss) and processed using Adobe Photoshop 7.0.
Goat anti mouse igg dylight 488
Manufactured by Thermo Fisher Scientific
Sourced in United States
Goat anti-mouse IgG-DyLight 488 is a secondary antibody conjugate designed for use in immunodetection techniques. It is composed of goat-derived antibodies specific to mouse immunoglobulin G (IgG) that are covalently linked to the DyLight 488 fluorescent dye.
Automatically generated - may contain errors
Lab products found in correlation
Irdye 680 goat anti rabbit igg, by LI COR (2 mentions)
Irdye 800 goat anti mouse igg, by LI COR (2 mentions)
Goat anti rabbit igg dylight 550, by Thermo Fisher Scientific (2 mentions)
Lsm 780, by Zeiss (1 mentions)
Streptomycin, by Thermo Fisher Scientific (1 mentions)
L glutamine, by Thermo Fisher Scientific (1 mentions)
Penicillin, by Thermo Fisher Scientific (1 mentions)
Hek293 cell, by American Type Culture Collection (1 mentions)
Acetylated tubulin, by Merck Group (1 mentions)
Pericentrin, by Fortrea (1 mentions)
Goat anti rabbit igg hrp, by Santa Cruz Biotechnology (1 mentions)
Mouse monoclonal anti flag antibody, by Merck Group (1 mentions)
Alexa fluor 488 goat anti mouse igg, by Thermo Fisher Scientific (1 mentions)
Goat anti mouse igg hrp, by Santa Cruz Biotechnology (1 mentions)
Lsm 510, by Zeiss (1 mentions)
Anti γ tubulin gtu 88, by Merck Group (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions)
A31571, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 647 donkey anti mouse igg, by Thermo Fisher Scientific (1 mentions)
A2547, by Merck Group (1 mentions)
7 protocols using goat anti mouse igg dylight 488
1
Immunofluorescence Analysis of Centrosomal Proteins
2
Yeast Protein Localization and Detection
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Saccharomyces cerevisiae strains used in this study are listed in Supplementary Table S2 . Strains were generated using standard cloning protocols. Anti-Kar2 polyclonal rabbit antibody and anti-Sec61 polyclonal rabbit antibody were gifts from Davis Ng (Temasek Life Sciences Laboratories, Singapore). Anti-HA mouse monoclonal antibody HA.11 (Covance), anti-Pgk1 mouse monoclonal antibody (Invitrogen), anti-GFP mouse monoclonal antibody (Roche) anti-tubulin mouse monoclonal antibody 12G10 (DHSB), anti-myc mouse monoclonal antibody (Invitrogen), anti-Flag mouse monoclonal antibody (Sigma), anti-LexA monoclonal mouse antibody (Santa Cruz Biotechnology) and anti-LexA polyclonal rabbit antibody (Abcam) were commercially purchased. Secondary antibodies goat anti-mouse IgG-DyLight 488 (Thermo Fisher, Waltham, MA), goat anti-rabbit IgG-DyLight 550 (Thermo Fisher), goat anti-mouse IgG-AlexaFluor488 (Invitrogen), goat anti-mouse IgG-HRP (Santa Cruz Biotechnology), goat anti-rabbit IgG-HRP (Santa Cruz), goat anti-mouse IgG-IRDye 800 (LI-COR Biosciences) and goat anti-rabbit IgG-IRDye 680 (LI-COR Biosciences) were commercially purchased.
3
Yeast Strain Preparation and Antibody Validation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
S. cerevisiae strains used in this study are listed in Table S7 . Strains were prepared using standard transformation protocols. Anti-HA mouse monoclonal antibodies HA.11 (MMS-101R-1000, Covance), anti-FLAG mouse M2 monoclonal antibody (F-1804, Sigma-Aldrich), and anti-Tub1 mouse monoclonal antibody (12G10, Developmental Studies Hybridoma Bank) were commercially purchased. Anti-Kar2 rabbit polyclonal was a gift from D. Ng (Temasek Life Sciences Laboratories, Singapore). Secondary antibodies goat anti-mouse IgG-DyLight 488 (35503, Thermo Fisher Scientific), goat anti-rabbit IgG DyLight 550 (84541, Thermo Fisher Scientific), goat anti-mouse IgG-IRDye 800 (926–32210, LI-COR Biosciences), and goat anti-rabbit IgG-IRDye 680 (926–68021, LI-COR Biosciences) were commercially purchased.
4
Immunofluorescence Analysis of Mouse Testes
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Fresh testes from laboratory mice of strain C57/Bl6 were minced in PBS containing 2% paraformaldehyde, 0.02% SDS, and 0.15% Triton X-100, and transferred onto superfrost slides. Cells were blocked for 1 h in PBS containing 1% BSA and 0.3% Triton X-100. Anti-CFAP52 (CSB-PA839781LA01HU; Wuhan Huamei Biotech Co., Ltd., Wuhan, China, Cusabio) was used as 1:100 dilution and incubated at 4 °C overnight. CFAP52 was detected using the secondary antibody goat anti-rabbit-IgG (H + L) Alexa Fluor R 555 (F[ab]2 fragment; #A21430, Life Technologies) used as 1:1,000 dilution. Additionally, the following first antibodies were used: anti-α-tubulin (DM1A, Calbiochem, #CP06), anti-γ-tubulin (GTU-88, Sigma-Aldrich, #T6557), anti-EB3 (EB3(7), Santa Cruz, sc-136405), anti-SUN3 and anti-SUN436 (link),50 (link), and detected using goat anti-mouse-IgG DyLight488 (Thermo Scientific, #35503) or goat anti-guinea pig-IgG Alexa Fluor 488 (Life Technologies, #A11073). DNA was counterstained with DAPI (4′,6-Diamidino-2-phenylindole; Sigma D-9542), and the acrosome was decorated with FITC-labelled peanut lectin (PL-FITC). Images were taken by confocal microscopy (LSM 510, Zeiss) and processed using Adobe Photoshop 7.0.
5
Immunofluorescence Staining of HEK-293 Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
HEK-293 cells were grown in poly-d -lysine–coated (Sigma-Aldrich) coverslips. After treatment, the cells were rinsed with PBS twice, fixed for 10 min in 4% paraformaldehyde, and permeabilized for 10 min with 0.1% Triton X-100 and 100-mM glycine in PBS. The cells were blocked for 60 min with PBT (PBS + 1.5% BSA and 0.1% Tween 20). Primary antibodies used were rabbit anti–TDP-43 (1:500; 10782-2-AP; Proteintech Group, Inc.), mouse anti–TIA-1 related protein (TIAR; 1:500; BD), and mouse anti–α-smooth muscle actin (1:100; A2547; Sigma-Aldrich). Secondary antibodies used were goat anti–mouse IgG, DyLight 488, goat anti–rabbit IgG, DyLight 594 (both 1:750; Thermo Fisher Scientific), and donkey anti–mouse IgG Alexa Fluor 647 (1:750; A31571; Life Technologies). Primary antibodies were diluted in PBT and incubated with the coverslip for 60 min. After washing the cells twice with PBS, they were incubated with the secondary antibody for 60 min in the dark. Cells were washed three times with PBS and mounted in mowiol containing DAPI.
6
Quantifying CP110 at Basal Bodies
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were transfected with p138NC::mRFP::FKBP and either pFRB::ECFP::Neurl4 or pFRB::ECFP::Hyls1, and dimerization was induced by incubation with rapamycin (#553210, 1 µM stock solution in DMSO, Millipore, Burlington, MA, USA). As a control, the cells were incubated with the same amount of DMSO, which was used as the solvent for rapamycin, for the indicated time. For quantification of CP110 at the basal bodies, the cells were incubated with anti-CP110 antibodies (#12780-1-AP, diluted 1:100, Proteintech, St. Leon-Rot, Germany), followed by goat anti-rabbit-IgG-AlexaFluor-647 (#A-21245, lot AB_2535813, Thermo Fisher Scientific, Waltham, MA, USA). The basal bodies were identified by the decoration of the primary cilium with anti-acetylated tubulin antibodies, followed by goat anti-mouse-IgG-Dylight 488 (#35503, Thermo Fisher Scientific, Waltham, MA, USA).
7
Immunofluorescence Assay for Bcl-2
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were treated with IC 50 /2 doses of drugs for 24 hr. Fixed HepG2 cells were blocked with bovine serum albumin (Capricorn, Ebsdorfergrund, Germany) and incubated with anti-human Bcl-2 antibody (1:100 dilution, Biocare med) overnight at 4 °C. Next day, the cells were rinsed with PBS three times and incubated with goat anti-mouse IgG DyLight 488 (1:200, Thermo Scientific, Waltham, MA, USA) for 90 min at 37 °C. After rinsing three times with PBS, the cells were counterstained with Hoechst 33258 (Thermo Scientific) for labeling the nuclei. Images were taken with a laser scanning confocal microscope (LSM 700, Zeiss).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!