Eight-well Nunc chamber slides (VWR, Darmstadt, Germany) were coated, washed, and blocked as described above. Wells were incubated with the serum of one NHD with a low SNEC ELISA value and the serum of one SLE patient with a high SNEC ELISA value (1:200 in PBS-T) or antibodies detecting histone H3 (mAb #34) and mouse anti-H3-K27me3 (mAb BT164). Goat anti-human IgG FITC (Jackson ImmunoResearch, Suffolk, UK), goat anti-mouse IgG Cy®5 (Jackson ImmunoResearch, Suffolk, UK), and goat anti-rabbit IgG Cy®5 (Jackson ImmunoResearch, Suffolk, UK) were added and incubated for 1 h in the dark. A control staining lacking a primary antibody was included. Finally, slides were washed with PBS and H2O, embedded in DAKO fluorescent mounting medium (Agilent Technologies, Santa Clara, CA, USA) and analyzed using the Eclipse Ni-U (Nikon Corporation, Tokyo, Japan).
Goat anti rabbit igg cy 5
Manufactured by Jackson ImmunoResearch
Sourced in United States
Goat anti-rabbit IgG Cy®5 is a secondary antibody conjugated with the fluorescent dye Cyanine 5 (Cy®5). It is designed to detect and visualize rabbit primary antibodies in various immunoassays and imaging applications.
Automatically generated - may contain errors
Lab products found in correlation
Goat anti mouse igg cy 5, by Jackson ImmunoResearch (4 mentions)
Cy3 goat anti rabbit igg, by Jackson ImmunoResearch (3 mentions)
Goat anti mouse igg alexa 488, by Jackson ImmunoResearch (2 mentions)
Goat anti mouse igg cy3, by Jackson ImmunoResearch (2 mentions)
Goat anti human igg fitc, by Jackson ImmunoResearch (1 mentions)
Dako fluorescent mounting medium, by Agilent Technologies (1 mentions)
Eclipse ni u, by Nikon (1 mentions)
A2066, by Merck Group (1 mentions)
Alexa 488 goat anti rat igg, by Jackson ImmunoResearch (1 mentions)
Hrp conjugated secondary antibody, by Jackson ImmunoResearch (1 mentions)
A0024, by Agilent Technologies (1 mentions)
Mouse anti myc tag, by Roche (1 mentions)
Mouse anti flag tag, by Merck Group (1 mentions)
Mouse anti rab27a, by Abcam (1 mentions)
Tcs sp5, by Leica (1 mentions)
Mouse anti mbp, by Abcam (1 mentions)
Immunol staining blocking buffer, by Beyotime (1 mentions)
E cadherin, by Cell Signaling Technology (1 mentions)
N cadherin, by Cell Signaling Technology (1 mentions)
Rat anti ha, by Roche (1 mentions)
6 protocols using goat anti rabbit igg cy 5
1
Fluorescent Immunostaining of Serum Antibodies
2
Comprehensive Immunofluorescence and Western Blot Antibodies
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The following primary antibodies were used for immunofluorescence microscopy (IFM) and/or immunoblotting (IB): isoform‐specific rabbit antiserum (AS) to P1c [6 ], rat monoclonal antibodies (mAbs) to α‐tubulin (clone YL1/2, SM2202P; Acris, Hiddenhausen, Germany), rabbit AS to tau (A‐0024; Dako, Glostrup, Denmark), mouse mAbs to α‐tubulin (clone B‐5‐1‐2, T5168; Sigma‐Aldrich), rabbit AS to GAPDH (G9545; Sigma‐Aldrich), mouse mAbs to GST (clone GST2, G1160; Sigma‐Aldrich), mouse mAbs to acetylated tubulin (clone 6‐11B‐1, T6793; Sigma‐Aldrich), rabbit AS to actin (A2066; Sigma‐Aldrich), rabbit AS to MAP1A/B [10 ], rabbit AS to MAP2 [10 ], rabbit AS to MAP1A light chain (LC) [11 ] and rabbit AS to MAP1B LC [12 ] (see also Table S1 ). For IFM, primary antibodies were used in combination with goat anti‐rat IgG Rhodamine red, goat anti‐rat IgG Alexa 488, goat anti‐mouse IgG Alexa 488, goat anti‐mouse IgG Cy5, goat anti‐rabbit IgG Alexa 488, goat anti‐rabbit IgG Cy5 (all from Jackson ImmunoResearch Laboratories, West Grove, PA, USA). For immunoblot analyses HRP‐conjugated secondary antibodies were used (Jackson ImmunoResearch Laboratories).
3
Immunostaining of SUMO Pathway Proteins
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Mouse anti-Gephyrin (1:1000, clones mAb7a, Synaptic Systems #147021), rabbit anti-SUMO-1 (1:250, Epitomics#1563-1), mouse anti-SUMO-1 (1:100, SantaCruz#sc-5308), rabbit anti-SUMO-2/3 (1:250, Cell signaling #4974), rabbit anti-SUMO-2/3 (1:250, Epitomics #2970-1), mouse anti-PIAS-3 (1:500, Sigma #P0117), rabbit anti-vGAT (1:2000, Synaptic Systems #131011); mouse anti-Myc tag (1:5000,Roche #11667149001), rabbit anti-Myc tag (1:5000, Cell Signaling #2278S), and mouse anti-FLAG tag (1:5000, Sigma Aldrich #F3165). All the secondary antibodies were from Jackson ImmunoResearch: Goat anti-Mouse Cy3 IgG (1:500, #115165), Goat anti-Mouse IgG Cy5 (1:500, #115175), Goat anti-Rabbit IgG Cy3 (1:500, #111165), and Goat anti-Rabbit IgG Cy5 (1:500, #111175).
4
Immunofluorescent Labeling of Myelin Proteins
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The culture medium was aspirated, and the cells were washed with PBS and then fixed with 4% paraformaldehyde (PFA) for 15 minutes at room temperature and methanol for 10 minutes at −20°C. The non-specific antibody-binding sites were blocked with Immunol Staining Blocking Buffer (Beyotime, Haimen, Jiangsu Province, China) for 1 hour at room temperature, and then the cultures were incubated with primary antibody overnight at 4°C. Antibodies to the following proteins were used at the indicated concentrations: mouse anti-Rab27a (1:100; Abcam, Cambridge, MA, USA), rabbitanti-Slp2-a (1:200; Protein Tech, Chicago, IL, USA), mouse anti-MBP (1:200, Abcam), chicken anti-P0 (1:50; Novus, Littleton, CO, USA), rabbit anti-MAG (1:200; Sigma), and mouse anti-NF (1:600; Sigma). After washing 3 times in PBS to remove excess primary antibody, the cultures were incubated for 1 hour at room temperature with their respective fluorescence-conjugated secondary antibodies (Jackson Immuno Research, West Grove, PA, USA): goat anti-rabbit IgG-Cy3 (H + L), goat anti-mouse IgG-Cy3 (H + L), goat anti-rabbit IgG-Cy5 (H + L), goat anti-mouse IgG-Alexa-488 (H + L), goat anti-rabbit IgG-Alexa-488 (H + L), and goat anti-chicken IgG (H + L) rhodamine, all at a dilution of 1:1,000. After a final PBS washing, the stainings were imaged with a confocal microscope (TCS SP5; Leica Microsystems, Wetzlar, Germany).
5
Immunofluorescence Analysis of Epithelial-Mesenchymal Transition
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Cells were fixed in 3.7% paraformaldehyde, washed twice with PBS after fixation, and then blocked in 2% FBS/2% BSA in PBS with 0.1% NP40 for 45 min. Cells were then treated with antibodies of E-CADHERIN (Cell Signaling Technology, 14,472) and N—CADHERIN (Cell Signaling Technology, 13,116) for 1 hour. After washed, samples were incubated with secondary antibodies in the same blocking buffer (Goat Anti-house IgG, Cy3, A22210, Abbkine, and Goat Anti-rabbit IgG, Cy5, 111–175–144, Jackson). After washed, DAPI solution was added to each sample and incubate for 15 min, wash twice. Mounting solution and coverslip were added. Samples were observed with fluorescence microscope.
6
Antibody Detection and Quantification
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The antbodies used are as follows: rabbit anti-STREP horseradish peroxidase (1:10,000, BioRad, 161-0380), Mouse anti-Gephyrin 3B11 (1:1,000, SySy, 147111), mouse anti-gephyrin mAb7a (1:2,000, SySy, 147021), rabbit anti-vGAT (1:3,000, SySy, 131003), mouse anti-vGAT (1:5,000, SySy, 131011), rat anti-HA (1:3,000, Roche), mouse anti-Myc (1:5,000, Roche, 11667149001), mouse anti-FLAG (1:5,000, Sigma-Aldrich, F3165), rabbit anti-GFP (1:5,000, Synaptic Systems, 132002), chicken anti-GFP (1:5,000, Chemicon, AB16901), mouse anti SUMO-1 (1:1,000, 21C7, Hybridoma bank), mouse anti SUMO2/3 (1:1,000, 8A2, Hybridoma Bank), rabbit anti-SUMO1 (1:250, Epitomics, 1563-1), rabbit anti-SUMO2/3 (1,250, Epitomics, 2970-1), rabbit anti pan-Acetylated Lysine antibody (1:1,000, Cell Signaling, 9441) and mouse anti-Ac-Lysine (AKL5C1) (1:2,000, Santa Cruz, Sc32268). Gephyrin anti-S270 phospho-antibody has been described earlier28 (link). The following secondary antibodies were used: goat anti-mouse IgG Cy3 (1:500, Jackson Immunoresearch, 115165), goat anti-mouse IgG Cy5 (1:500, Jackson Immunoresearch, 115175), goat anti-rabbit IgG Cy3 (1:500, Jackson Immunoresearch, 111165), goat anti-rabbit IgG Cy5 (1:500, Jackson Immunoresearch, 111175), goat anti-mouse IgG alkaline phosphatase (AP) (1:30,000, Sigma-Aldrich, A3562) and goat anti-rabbit IgG AP (1:30,000, Sigma-Aldrich, A3687).
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