Alexa 555 conjugated anti rabbit igg

Manufactured by Thermo Fisher Scientific

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Alexa Fluor 555-conjugated anti-rabbit IgG is a secondary antibody that binds to rabbit primary antibodies. It is used to detect and visualize rabbit-derived target proteins in various immunoassays, such as Western blotting, immunohistochemistry, and flow cytometry.

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Lab products found in correlation

Alexa 488 conjugated anti mouse igg, by Thermo Fisher Scientific (4 mentions) Axioplan epifluorescence microscope, by Zeiss (2 mentions) Anti tub, by Merck Group (2 mentions) Fitc conjugated anti mouse igg igm, by Jackson ImmunoResearch (2 mentions) Vectashield medium h 1200 with dapi, by Vector Laboratories (2 mentions) Streptavidin alexa 488, by Thermo Fisher Scientific (1 mentions) Axiocam hrc, by Zeiss (1 mentions) Axiovision, by Zeiss (1 mentions) Biotinyl tyramide solution, by PerkinElmer (1 mentions) Eclipse e800m, by Nikon (1 mentions) Biotinylated goat anti rabbit igg, by PerkinElmer (1 mentions) 4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions) Evos fl fluorescence microscope, by Thermo Fisher Scientific (1 mentions) Anti phospho histone h2ax s139, by Cell Signaling Technology (1 mentions) Lsm 710 confocal microscope, by Zeiss (1 mentions) Phosphatase inhibitor cocktail, by Roche (1 mentions) Anti γh2ax, by Merck Group (1 mentions) Anti flag, by Merck Group (1 mentions) Anti pcna, by Santa Cruz Biotechnology (1 mentions) Protease inhibitor cocktail, by Merck Group (1 mentions)

Spelling variants of this product

Alexa 555 (299 citations) Alexa Fluor 555-conjugated anti-rabbit IgG (35 citations) Alexa Fluor 555-conjugated goat anti-rabbit IgG secondary antibody (7 citations) IgG Rabbit (4 citations) Alexa Fluor 555 conjugated anti-rabbit IgG secondary antibody (4 citations) Alexa Fluor 555-conjugated goat anti-rabbit IgG (H + L) (4 citations) Goat anti-rabbit IgG conjugated with Alexa Fluor 555 (4 citations) Alexa 555 anti-rabbit IgG (2 citations) Alexa Fluor® 555 dye-conjugated anti-rabbit IgG (2 citations) Goat anti-rabbit IgG conjugated to Alexa 555 (2 citations)

9 protocols using alexa 555 conjugated anti rabbit igg

Kisspeptin and NKB Double-Labeling Immunohistochemistry

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Every 6th section of the diencephalon was processed for kisspeptin/NKB double-labeling fluorescence immunohistochemistry using the tyramide signal amplification (TSA) method [8 (link), 33 (link)]. Sections were sequentially incubated with 10% NGS in PBST-BSA for 1 h, anti-NKB polyclonal antibody (1:6,000) in PBST-BSA containing 2% NGS at 4°C for 72 h, biotinylated goat anti-rabbit IgG
(1:200 in PBST-BSA containing 2% NGS) for 3 h, and avidin-biotin complex (2 µl/ml PBST) for 1 h. After being soaked in the TSA blocking solution for 30 min, the sections were reacted with biotinyl tyramide solution (1:200,
PerkinElmer, Waltham, MA, USA) for 10 min, and streptavidin-Alexa 488 (1:200, Invitrogen, Carlsbad, CA, USA) in PBST for 1 h. They were then incubated with gC2 (1:2,000) for 48 h at 4°C and Alexa-555 conjugated anti-rabbit IgG
(1:200, Invitrogen) for 3 h. Kisspeptin/Dyn double-labeling immunohistochemistry was conducted using the same protocol and the anti-Dyn polyclonal antibody (1:20,000). Sections were mounted on gelatin-coated slides and
cover-slipped with a water-soluble mounting medium (Vector Laboratories). The sections were observed under a microscope (ECLIPSE E800M, Nikon, Tokyo, Japan) equipped with a charge-coupled device camera (AxioCam HRc, Carl Zeiss,
Jena, Germany). The two fluorescent images were merged using computer software (AxioVision, Carl Zeiss).

OKAMURA H., YAMAMURA T, & WAKABAYASHI Y. (2017). Mapping of KNDy neurons and immunohistochemical analysis of the interaction between KNDy and substance P neural systems in goat. The Journal of Reproduction and Development, 63(6), 571-580.

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Quantifying DNA Damage Foci in Cells

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Cells were seeded in 48-well plates and incubated for 24 hours before treatment with etoposide (6 μM) for another 24 hours. After treatment, cells were washed twice with ice-cold PBS, fixed in a solution containing 2% paraformaldehyde and 1% sucrose for 15 minutes at room temperature, and permeabilized with ice-cold methanol and acetic acid (1:1) for 20 minutes at −20°C. Cells were washed in PBS and then incubated in a blocking buffer (10% horse serum) for 60 minutes. For ionizing irradiation, cells were fixed and processed at different time points after X-irradiation.
Incubation with rabbit polyclonal anti-53BP1 antibody (1:200, Santa Cruz, sc22760) or anti-phospho-Histone H2AX-S139 (1:200, Cell Signaling, 9718) was carried out at 4°C overnight, followed by incubation with Alexa 555–conjugated anti-rabbit IgG (1:500; Invitrogen) for 1 hour at room temperature in the dark. Nuclei were counterstained with DAPI (0.2 μg/mL, Sigma-Aldrich). Immunofluorescence was examined and pictures taken using the EVOS-FL fluorescence microscope (ThermoScientific). Nuclei with >5 foci were counted. Percentage of foci⁺ cells = number of foci⁺ cells divided by total number of cells counted x 100%.

Kim H., Lin Q, & Yun Z. (2018). The hypoxic tumor microenvironment in vivo selects the tumor cells with increased survival against genotoxic stresses. Cancer letters, 431, 142-149.

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Confocal Microscopy-Based Colocalization Analysis

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U2OS cells were labeled with EdU for 30 min, rinsed in phosphate buffered saline (PBS) pH 7.4 and then pre-extracted in PBS-T buffer (0.2% Triton X-100 in 1× PBS, phenylmethylsulphonylfluoride [PMSF], protease inhibitor cocktail [Sigma, P8340] and phosphatase inhibitor cocktail [Roche, P4906845001]) for 5 min on ice, followed by 2.0% paraformaldehyde (PFA). Primary antibody staining was performed as follows: anti-FLAG (1:1000, Sigma, F1804), anti-CUL4A (1:200, Abcam, ab92554), anti-γH2AX (1:500, Millipore, 05-636), anti-pRPA (1:500, Bethyl labs, A300-245A) and anti-PCNA (1:200, Santa Cruz, sc-7907). Secondary antibody staining was performed as follows: Alexa 488 conjugated anti-mouse IgG and Alexa 555 conjugated anti-rabbit IgG (1:500, Thermo Fisher Scientific, A11029 and A21428). EdU was detected by Click-iT assay kit according to the manufacturer’s protocol. A Zeiss LSM710 confocal microscope was used and Pearson’s correlation coefficient (R, or Rcoloc) that is the covariance of the two variables divided by the product of their standard deviations was used for colocalization analysis and calculated using the colocalization Plugin of the FIJI-ImageJ software (https://imagej.nih.gov/ij/index.html). Detailed results of the colocalization analyses are presented in Supplementary Dataset.

Jang S.M., Zhang Y., Utani K., Fu H., Redon C.E., Marks A.B., Smith O.K., Redmond C.J., Baris A.M., Tulchinsky D.A, & Aladjem M.I. (2018). The replication initiation determinant protein (RepID) modulates replication by recruiting CUL4 to chromatin. Nature Communications, 9, 2782.

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Zebrafish Brain Immunohistochemistry Protocol

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Zebrafish larvae were sacrificed with 0.04% tricaine and fixed in 4% paraformaldehyde at 4°C overnight. Following Genes to Cells HIGUCHI rinses with a phosphate-buffered saline containing 0.05% Tween 20 (PBST), the brain was manually isolated and blocked with PBST containing 2% normal goat serum (NGS) for 1 hr at room temperature. Samples were then incubated with the primary antibodies in PBST with 2% NGS at 4°C overnight. Primary antibodies used in this study include those against PCNA (Merck-Millipore) and phosphohistone H3 (Merck-Millipore). Samples were rinsed three times with PBST and then incubated with the secondary antibodies in PBST with 2% NGS at room temperature for 2 hr. Secondary antibodies used in this study include Alexa 488-conjugated anti-mouse IgG (Thermo Fisher Scientific) and Alexa 555-conjugated anti-rabbit IgG (Thermo Fisher Scientific).

Higuchi M. (2020). Maternal stress suppresses cell proliferation in the forebrain of zebrafish larvae. Genes to cells : devoted to molecular & cellular mechanisms, 25(5).

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Immunofluorescent Staining of Phosphorylated Myosin Light Chain

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Cells were fixed with 3.7% formaldehyde in PBS for 20 min, followed by permeabilization by 5 min incubation in 0.05% Triton X-100-containing PBS. Samples were soaked for 30 min in Can Get Signal immunostain (solution A) (Toyobo, Japan), and then incubated with ppMLC antibody (1:50 dilution; Cell Signaling Technology #3674) in Can Get Signal immunostain (solution A) for 1 h at room temperature. Next, the cells were washed 3 times with PBS, and then incubated for 1 h at room temperature with Alexa 555-conjugated anti-rabbit IgG (1:100 dilution; ThermoFisher) in Can Get Signal immunostain (solution A). Finally, the cells were washed 3 times with PBS and subjected to fluorescence imaging.

Yamamoto K., Miura H., Ishida M., Sawai S., Kondo Y., & Aoki K. (2021). Optogenetic relaxation of actomyosin contractility uncovers mechanistic roles of cortical tension during cytokinesis.

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Immunostaining of Drosophila Testes

Cited 2 times

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Fixation and F-actin staining of Drosophila testes were performed as described in [14 (link)]. Fixation for the other immunostainings was performed as previously described [15 (link),16 (link)]. The primary antibodies and the dilutions (in PBS) used were as follows: Anti-tub (1:1000) (Sigma-Aldrich, St. Louis, MO, USA), anti-Pav (1:100) [17 (link),18 (link)], anti-Spd2 (1:5000) [19 (link)], anti-Feo (1:50) [20 (link)], anti-HTS (IBI) (1:5) (Hybridoma Bank, The University of Iowa, IA, USA) [21 (link)], anti-anillin (1:1000) [22 (link)]. The secondary antibody incubation was performed using both the FITC-conjugated anti-mouse IgG+IgM (1:20 in PBS; Jackson ImmunoResearch Laboratories, Cambridge, UK) and Alexa 555-conjugated anti-rabbit IgG (1:300 in PBS; Molecular Probes, Eugene, OR, USA) for 2 h at room temperature. Slides were then mounted in Vectashield medium H-1200 with DAPI (Vector Laboratories, Burlingame, CA, USA) to stain DNA and reduce fluorescence fading.
Slides with mitotic chromosome preparations and fixed testes were analyzed using a Zeiss Axioplan epifluorescence microscope (Carl Zeiss, Obezkochen, Germany), equipped with a cooled CCD camera (Photometrics, Woburn, MA, USA). Gray-scale digital images were collected separately, converted to Photoshop format, pseudocolored, and merged.

Di Giorgio M.L., Morciano P., Bucciarelli E., Porrazzo A., Cipressa F., Saraniero S., Manzi D., Rong Y.S, & Cenci G. (2020). The Drosophila Citrate Lyase Is Required for Cell Division during Spermatogenesis. Cells, 9(1), 206.

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Gatastatin Effects on MG132-Treated Spindles

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To observe the effect of gatastatin on MG132-treated metaphase-like spindle using 3D-SIM (three-dimensional structural illumination microscopy), HeLa cells were arrested at metaphase by sequential treatment with STLC and MG132. After treatment with 1% DMSO or 30 μM of gatastatin for 2 h (combination treatment with MG132 and gatastatin), cells were fixed with cold MeOH (−20 °C). Cells were blocked with PBS containing 0.5% bovine serum albumin and then were incubated with anti-α-tubulin (1:500 dilution, Sigma-Aldrich Cat# T9026) and γ-tubulin (1:500 dilution, antibody against C-EYHAATRPDYISWGTQDK peptide of γ-tubulin43 (link)) antibodies. Secondary antibodies were Alexa⁴⁸⁸-conjugated anti-mouse IgG (1:500 dilution, Molecular Probes) and Alexa555-conjugated anti-rabbit IgG (1:500 dilution, Molecular Probes). DNA was stained with 0.5 μg ml⁻¹ Hoechst 33342 for 30 min. Finally, cells were washed four times with PBS and mounted in Prolong Gold Antifade (Molecular Probes, #P36930). Spindle structure was observed under a 3D-SIM-Nikon Ti inverted microscope.

Chinen T., Liu P., Shioda S., Pagel J., Cerikan B., Lin T.C., Gruss O., Hayashi Y., Takeno H., Shima T., Okada Y., Hayakawa I., Hayashi Y., Kigoshi H., Usui T, & Schiebel E. (2015). The γ-tubulin-specific inhibitor gatastatin reveals temporal requirements of microtubule nucleation during the cell cycle. Nature Communications, 6, 8722.

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Immunofluorescence Analysis of Neuronal Markers

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Cells were fixed with 4% (w/v) paraformaldehyde for 15 min at room temperature and permeabilized using 0.2% Triton X-100 for 5 min at room temperature. The cells were then blocked using 5% (w/v) BSA in PBS and incubated with an anti-βIII-tubulin antibody (Abcam), an anti-LIS1 antibody, an anti-GSK-3β antibody (BD Biosciences), an anti-pS9-GSK-3β antibody (Cell Signaling Technology, CST, Danvers, USA), anti-pY216-GSK-3β antibody (BD Biosciences), and anti-active Caspase-3 antibody (Abcam) followed by incubation with Alexa 488-conjugated anti-mouse IgG, Alexa 555-conjugated anti-rabbit IgG, and/or Alexa 647-conjugated anti-rabbit IgG (Molecular Probes, Eugene, OR, USA) as appropriate. Actin filament was stained by Rhodamine Phalloidin Conjugate (Thermo Fisher Sceintific). Nuclei were counterstained using 100 nM 4′,6-diamidino-2-phenylindole (DAPI). Each incubation was performed for 1 h at room temperature. Slides were mounted in FluorSave Reagent (345789, Calbiochem, La Jolla, CA, USA). Immunofluorescence was measured under a laser scanning confocal microscope (TCS-SP5, Leica, or LSM 700, Carl Zeiss) under the control of accessory software (LAS AF, Leica, or ZEN 2012, Carl Zeiss). Nuclei were labeled with DAPI (Thermo Fisher Scientific).

Kumamoto K., Iguchi T., Ishida R., Uemura T., Sato M, & Hirotsune S. (2017). Developmental downregulation of LIS1 expression limits axonal extension and allows axon pruning. Biology Open, 6(7), 1041-1055.

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Imaging Drosophila Testis Morphology

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Fixation and staining were performed on dissected third-instar larvae testes as previously described [8 (link)]. The primary antibodies and the dilutions used were as follows: Anti-tub (1:1000) (Sigma-Aldrich, St. Louis, MO, USA), anti-DmATPCL (1:100) [14 (link)], anti-Lava lamp (1:300) [16 (link)]. The secondary antibody incubation was performed using FITC-conjugated anti-mouse IgG + IgM (1:20; Jackson ImmunoResearch Laboratories, Cambridge, UK), Alexa 555-conjugated anti-rabbit IgG (1:300 in PBS; Molecular Probes, Eugene, OR, USA) and Alexa Fluor 488-conjugated anti-guinea pig IgG (1:300 in PBS; Jackson laboratories) for 2 h at room temperature. Slides were then mounted in Vectashield medium H-1200 with DAPI (Vector Laboratories, Burlingame, CA, USA) to stain DNA and reduce fluorescence fading. Slides were analyzed using a Zeiss Axioplan epifluorescence microscope (Carl Zeiss, Obezkochen, Germany), equipped with a cooled CCD camera (Photometrics, Woburn, MA, USA). Gray-scale digital images were collected separately, converted to Photoshop format, pseudo-colored, and merged. Quantification of Lva spot was carried out using the ImageJ software. Box plots were obtained with Python 3.3.

Morciano P., Di Giorgio M.L., Tullo L, & Cenci G. (2021). The Organization of the Golgi Structures during Drosophila Male Meiosis Requires the Citrate Lyase ATPCL. International Journal of Molecular Sciences, 22(11), 5745.

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