TDCs were seeded at a cell density of 5 × 104 cells/well and incubated at 37 °C at 5% CO2. After 24 h incubation, cells were fixed at 37 °C for 10 min with 4% formaldehyde (pH 7.4). They were then washed twice with PBS and permeabilized at room temperature for 10 min. Next, cells were blocked in blocking solution (2% bovine serum albumin in PBS) for 1 h at room temperature, incubated overnight with primary antibodies diluted in blocking solution at 4 °C, washed three times for 5 min with PBS, incubated with secondary antibody at RT for 2 h, and washed four times for 5 min with PBS. The primary antibodies used for immunostaining and their dilutions were as follows: rabbit anti-tenomodulin (1:50, Abcam, Cambridge, UK), mouse anti-CD44 (1:50, BD Biosciences), mouse anti-90(1:50, BD Biosciences). The secondary antibodies used were Alexa Fluor 488-conjugated goat anti-mouse IgG (1:100, Thermo, Waltham, US) and Alexa Fluor 555-conjugated goat anti-rabbit IgG (1:100, Thermo).
Alexa fluor 555 conjugated goat anti rabbit igg
Manufactured by Thermo Fisher Scientific
Sourced in United States, United Kingdom
Alexa Fluor 555-conjugated goat anti-rabbit IgG is a secondary antibody used for detection and visualization in immunoassays and other applications. It is a conjugate of a goat-derived antibody specific for rabbit immunoglobulin G (IgG) and the Alexa Fluor 555 fluorescent dye.
Automatically generated - may contain errors
Lab products found in correlation
Alexa fluor 488 conjugated goat anti mouse igg, by Thermo Fisher Scientific (11 mentions)
Alexa fluor 555 conjugated goat anti mouse igg, by Thermo Fisher Scientific (3 mentions)
Lsm 780 confocal microscope, by Zeiss (3 mentions)
Lsm 710 confocal microscope, by Zeiss (3 mentions)
Hoechst 33258, by Thermo Fisher Scientific (2 mentions)
Ab68538, by Abcam (2 mentions)
Cm3050, by Leica (2 mentions)
Mouse monoclonal anti gapdh, by Merck Group (2 mentions)
Anti ddx5, by Santa Cruz Biotechnology (2 mentions)
Anti akap95, by Santa Cruz Biotechnology (2 mentions)
Anti akap95, by Fortis Life Sciences (2 mentions)
Mouse anti srsf2 antibody, by Abcam (2 mentions)
Mouse monoclonal anti hnrnp m, by Santa Cruz Biotechnology (2 mentions)
Anti cyclin a, by Santa Cruz Biotechnology (2 mentions)
Lsm 780, by Zeiss (2 mentions)
Rabbit anti zo 1, by Thermo Fisher Scientific (2 mentions)
Alexa fluor 488 conjugated goat anti rabbit igg, by Thermo Fisher Scientific (2 mentions)
4 6 diamidino 2 phenylindole dapi, by Abcam (2 mentions)
Cellsense software, by Olympus (2 mentions)
Rabbit anti tenomodulin, by Abcam (1 mentions)
35 protocols using alexa fluor 555 conjugated goat anti rabbit igg
1
Immunofluorescence Staining of Tenomodulin, CD44, and CD90
2
Molecular Pathway Regulation in Cancer
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PLSCR1 shRNA and STAT3 shRNA were purchased from Sigma-Aldrich (St Louis, MO). Human PLSCR1, STAT3, and EGFR were amplified from the MDA-MB231 cDNA and subcloned into pLenti6.3⁄V5, pLVX, and pCMV, respectively.
Recombinant human EGF protein, human interleukin-6 (hIL-6), and antibodies against PLSCR1 and ALDH1 were purchased from Abcam (Carlsbad, CA). Antibodies against STAT1, STAT3, Phospho- STAT1 (Tyr701) and Phospho-STAT3 (Tyr705) were acquired from Cell Signaling Technology (Danvers, MA). Antibodies against phospho-Tyr were obtained from Santa Cruz (Dallas, TX). 6x-His Tag monoclonal antibody and Alexa Fluor 555-conjugated goat anti-rabbit IgG were purchased from Thermo Fisher Scientific. Antibodies for Flag, Myc, β-Tubulin, LaminB and Anti-FLAG Magnetic Beads were obtained from Sigma-Aldrich (St. Louis, MO).
Recombinant human EGF protein, human interleukin-6 (hIL-6), and antibodies against PLSCR1 and ALDH1 were purchased from Abcam (Carlsbad, CA). Antibodies against STAT1, STAT3, Phospho- STAT1 (Tyr701) and Phospho-STAT3 (Tyr705) were acquired from Cell Signaling Technology (Danvers, MA). Antibodies against phospho-Tyr were obtained from Santa Cruz (Dallas, TX). 6x-His Tag monoclonal antibody and Alexa Fluor 555-conjugated goat anti-rabbit IgG were purchased from Thermo Fisher Scientific. Antibodies for Flag, Myc, β-Tubulin, LaminB and Anti-FLAG Magnetic Beads were obtained from Sigma-Aldrich (St. Louis, MO).
3
Immunofluorescent Analysis of Bone Matrix Proteins
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Antigen retrieval was performed via incubation of specimens in a solution of 0.5 M acetic acid (Sigma-Aldrich) with 0.1% pepsin (Nacalai Tesque) at 37°C for 1 hour in a humid chamber prior to immunofluorescent staining. After washing, nonspecific binding was blocked for 60 minutes using 2% BSA (Wako Pure Chemical), 0.1% Tween 20 (Sigma-Aldrich), and 0.01% Triton-X (Wako Pure Chemical) prior to incubation in the primary antibody at 4°C overnight. After washing, the specimens were incubated with the secondary antibody for 1 hour at room temperature. Subsequently, fluorescence was detected by confocal microscopy (LSM780, Zeiss). The primary antibodies used in this study were anti-type I collagen monoclonal antibody (NB600-450: 1/50, Novus Biologicals, Littleton, CO, USA), anti-osteocalcin polyclonal antibody (FL-95: 1/50, Santa Cruz Biotechnology), and control IgG (normal mouse IgG (sc-2025): 1/50 or rabbit IgG (sc-2027): 1/50, Santa Cruz Biotechnology). Secondary antibodies were Alexa Fluor 488-conjugated goat anti-mouse IgG (1/500, Molecular Probes, Thermo Fisher Scientific) or Alexa Fluor 555-conjugated goat anti-rabbit IgG (1/500, Thermo Fisher Scientific). Nuclear staining was performed using Hoechst 33258 (1/500, Thermo Fisher Scientific).
4
Immunohistochemistry of Lung Tissues for P. aeruginosa Detection
5
Npba Immunofluorescence in ptger4b Mice
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Whole brains of ptger4b+/+ and ptger4b−/− females were fixed in 4% PFA, embedded in paraffin, and cut into 10 μm coronal sections. After blocking with phosphate-buffered saline (PBS) containing 2% normal goat serum, sections were incubated overnight at 4 °C with the anti-Npba antibody described above13 (link) diluted 1:1000 in PBS containing 2% normal goat serum, 0.1% bovine serum albumin, and 0.02% keyhole limpet hemocyanin. The sections were then incubated overnight at 4 °C with Alexa Fluor 555-conjugated goat anti-rabbit IgG (RRID: AB_2535849; Thermo Fisher Scientific) and DAPI diluted 1:1000 in PBS. Images were captured as described above, using excitation and emission wavelengths of 405 nm and 410–480 nm for DAPI and 552 nm and 562–700 nm for Alexa Fluor 555. The immunofluorescence intensity of Npba was measured using LAS X software (ver. 3.7.4; Leica Microsystems).
6
Immunostaining of Tight Junction Proteins
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Two or eighteen hours after the shockwave application, the rats were sacrificed and the brains were harvested and fixed with 10% formalin at room temperature overnight. Samples were then embedded in paraffin and serial 7-μm transverse sections around the shockwave treatment site were mounted on slides. The sections were deparaffinized, rehydrated, antigen retrieved (120℃, 10 min) and washed in TBS, followed by washing with TBS containing 0.025% Triton X-100 for 10 min. The sections were blocked with 10% newborn calf serum (NCS) and 1% BSA in TBS for 2 h. The sections were incubated with primary antibody overnight at 4℃. After washing with TBS, the samples were incubated with the secondary antibody for 2 h at RT, washed with TBS and mounted with EverBrite™ Hardset Mounting Medium containing DAPI to label the nuclei (Biotium). Slides were viewed, and images were captured with LSM780 confocal microscope (Zeiss, Jena, Germany). The primary antibodies used for immunostaining and their dilutions were as follows: rabbit anti-ZO1 (1:50, Thermo), mouse anti-occludin (1:50, Thermo), rabbit TRPV4 (1:50, Thermo). The secondary antibodies used were Alexa Fluor 488-conjugated goat anti-mouse IgG (1:100, Thermo) and Alexa Fluor 555-conjugated goat anti-rabbit IgG (1:100, Thermo).
7
Immunofluorescence analysis of sciatic nerve
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Sciatic nerves (n = 3/genotype) were rapidly dissected into 4% PFA and incubated for 30 min at 4℃. The nerve was washed in three changes of PBS and moved to 30% sucrose for 1 h at 4℃. Nerves were embedded in OCT mount medium and 10 µm longitudinal sections collected onto APES coated slides. Slides were pre‐treated with 100% EtOH at −20℃ for 10 min, thoroughly washed in PBS, before application of a blocking solution (0.3% Triton +10% normal goat serum) for 1 h at 4℃. Slides were then incubated overnight at 4℃ with either one of the following combinations of primary antibodies: rabbit anti‐voltage‐gated potassium channel (Kv1.1, Alomone Laboratories #APC‐009; RRID:AB_2040144; 1:200) plus mouse anti‐Caspr (Antibodies incorporated #75‐001; RRID:AB_ 2083496; 1:300); rabbit anti‐pan neurofascin (anti‐pan‐NFasc; gifted from Professor Brophy, University of Edinburgh, UK; 1:1000) plus mouse anti‐pan voltage‐gated sodium channel (pNav; Sigma‐Aldrich #8809; RRID:AB_477552; 1:100). Following washes in PBS, slides were incubated with secondary antibodies prepared in PBS plus 1% NGS for 2 h at R.T. as follows: Alexa Fluor 555‐conjugated goat anti‐rabbit IgG (Thermo Fisher Scientific Cat# A‐21429, RRID:AB_2535850; 1:500); Alexa Fluor 647‐conjugated goat anti‐mouse IgG1 antibody (Thermo Fisher Scientific Cat# A‐21240, RRID:AB_2535809; 1:500). After PBS washes, slides were mounted in Citifluor.
8
Immunostaining of Myelinated Nerve Fibers
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Sciatic nerves were rapidly dissected and desheathed in oxygenated (95% O2 and 5% CO2) physiological Ringer's solution containing the following (in mM): NaCl, 129; KCl, 3; NaH2PO4, 1.2; CaCl2, 2.4; MgSO4 1.3; HEPES, 3; NaHCO3, 20 and glucose, 10. Nerves were incubated for 1 h at 4℃ in 100µg/ml anti‐GM1 IgG3 antibody that has been generated as previously described (Boffey et al., 2005 (link); Townson et al., 2007 (link)). Nerves were washed and fixed in 4% PFA for 30 min at 4℃, then washed in three 10 min changes of PBS, 0.1 M glycine and PBS. Sciatic nerves were gently teased out into single fibres onto APES coated slides. Slides were incubated overnight at 4℃ in blocking solution (0.3% Triton +3% normal goat serum) with rabbit anti‐gliomedin antibody (Gldn, Abcam #ab24483, RRID:AB_2111616; 1:100). Nerves were washed with PBS followed by application of Alexa Fluor 488‐conjugated goat anti‐mouse IgG3 (Thermo Fisher Scientific Cat# A‐21151, RRID:AB_2535784; 1:500) and Alexa Fluor 555‐conjugated goat anti‐rabbit IgG (Thermo Fisher Scientific Cat# A‐21429, RRID:AB_2535850; 1:500) in PBS with 3% NGS for 2h at R.T. Slides were washed in PBS and mounted in Citifluor.
9
Immunoblotting for AKAP95 and Transcription Factors
10
Antibody Sources for Tau Protein Analysis
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Anti-tau (rabbit) antibody (catalog no. SC1996-R, lot no. B1213) was from Santa Cruz Biochemistry. Anti-tau pSer396 (rabbit) antibody (catalog no. BS4196, lot no. CJ36131) was from Bioworld Technology. Anti-FMRP (mouse) antibody (catalog no. MAB2160, lot no. 2137991, clone 1C3) was from Millipore. Rabbit anti-Staufen1 (catalog no. ab73478, Gr21579-1), mouse anti-Pur α (catalog no. ab77734, lot no. GR98153-2), and rabbit anti-YB-1 (catalog no. ab76149, GR221265-24, clone EP2708Y) antibodies were from Abcam. Anti-ZBP1/IMP1 (mouse) antibody (catalog no. RN001M, lot no. 001, clone 6H6) was from MBL Life Science. Alexa Fluor 555-conjugated goat anti-rabbit IgG (catalog no. A21428, lot no. 1937183) was purchased from Thermo Fisher Scientific. Horseradish peroxidase (HRP)-linked anti-rabbit IgG (donkey) (catalog no. NA934V. lot no. 377022) was from GE Healthcare Life Science.
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