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Fitc or rhodamine conjugated secondary antibodies

Manufactured by Jackson ImmunoResearch

FITC- or rhodamine-conjugated secondary antibodies are laboratory reagents used to detect and visualize primary antibodies in various immunoassays. These conjugated secondary antibodies bind to the Fc region of primary antibodies, allowing for the detection and localization of target antigens.

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4 protocols using fitc or rhodamine conjugated secondary antibodies

1

Genetically Engineered Mouse Models for Breast Cancer

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BRCA1f/f and Tg (MMTV-Cre) 4Mam mice were obtained from the NCI Mouse Repository and JAX lab, respectively [46 (link), 47 (link)]. The generation of p16−/−, p18−/−, BRCA1+/−, BRCA1MGKO(BRCA1f/f;MMTV-Cre or BRCA1f/−;MMTV-Cre) mice was described previously [31 (link), 48 (link)–50 (link)]. The Institutional Animal Care and Use Committee at the University of Miami and Shenzhen University approved all animal procedures. Histopathology and immunohistochemistry (IHC) were performed as described previously [11 (link), 28 (link), 31 (link)]. The primary antibodies used were Ck14, cleaved caspase 3 (Thermal Scientific), PDGFRβ, phosphorylated PKCα (p-PKCα), phosphorylated FRA1 (p-FRA1), FRA1 (Cell signaling), ERα, BRCA1 (Santa Cruz), E-cadherin (E-cad), PKCα (BD Biosciences), and Vimentin (Vim) (Abcam). Immunocomplexes were detected by using the Vectastain ABC DAB kit according to the manufacturer’s instructions (Vector Laboratories) or by using FITC- or rhodamine-conjugated secondary antibodies (Jackson Immunoresearch).
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2

Genetically Modified Mouse Models for Breast Cancer

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The generation of p18mt (p18−/− and p18+/−), p18mt;Gata3+/− (p18−/−;Gata3+/− and p18+/−;Gata3+/−), and p18mt;Brca1+/− (p18−/−;Brca1+/− and p18+/−;Brca1+/−) mice has been previously described [15 (link), 16 (link)]. MMTV-PyMT and NCG were purchased from GemPharmatech (Nanjing, China), and NSG mice were purchased from Jackson Laboratory (Maine, USA). The Institutional Animal Care and Use Committee at the University of Miami and Shenzhen University approved all animal procedures. Animals were housed in a specific pathogen-free environment. The investigators were not blinded to genotype allocation during experiments and outcome assessment. No randomization method was used as mice were segregated into groups based on genotype. Histopathology, immunohistochemistry (IHC), and immunofluorescence staining (IF) were performed as previously described [14 (link), 20 (link), 36 (link)]. The primary antibodies used were GATA3, FRA1, c-FOS, SNAIL, TWIST, E-cadherin (E-cad), Ki67, Fibronectin (Fn), Vimentin (Vim) (Cell Signaling). Immunocomplexes were detected using the Vectastain ABC alkaline phosphatase kit according to the manufacturer’s instructions (Vector Laboratories), or using FITC- or rhodamine-conjugated secondary antibodies (Jackson Immunoresearch).
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3

Genetically Engineered Mouse Models for Cancer Research

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The generation of p18-/-, p18+/-, GATA3+/-, p18+/-;GATA3+/-, and p18-/-;GATA3+/- mice was previously described 17 (link), 27 (link), 41 (link), 44 (link). NOD-Prkdcem26Cd52Il2rgem26Cd22/Nju (NCG) and FVB/NJGpt-Tg(MMTV-PyMT)/Gpt were purchased from GemPharmatech (Nanjing, China). The Institutional Animal Care and Use Committee at the University of Miami and Shenzhen University approved all animal procedures. Histopathology and immunohistochemistry (IHC) were performed as previously described 17 (link), 27 (link), 44 (link). The primary antibodies used were: E-cadherin (E-Cad) (BD Biosciences), Ck5 (Covance), Ck8 (American Research Products), Ck14 (Thermal Scientific), eGFP (GeneTex), GATA3 (Santa Cruz), SMA (Cell Signaling), and Ki67 (Abcam). Immunocomplexes were detected using the Vectastain ABC alkaline phosphatase kit according to the manufacturer's instructions (Vector Laboratories), or using FITC- or rhodamine-conjugated secondary antibodies (Jackson Immunoresearch). IHC results were quantified using H-score as previously described 45 (link), 46 (link).
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4

Immunofluorescence Imaging of Transfected HeLa Cells

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HeLa cells were grown on coverslips and transfected with the indicated plasmid. At 24 h after transfection, cells were washed once with PBS and fixed in 4% paraformaldehyde in PBS. Subsequently, cells were permeabilized with 0.2% Triton X-100 and treated for 30 min at room temperature with 10% BSA in PBS, followed by incubation with primary antibody for 1 h. Primary anti-Flag or anti-MAVS antibodies (Santa Cruz Biotechnology, Santa Cruz, CA) was used, followed by incubation with FITC- or Rhodamine-conjugated secondary antibodies (Jackson Immuno Research Laboratories, West Grove, PA), and 4-,6-diamidino-2-phenylindole (DAPI)was used to stain the nuclei. The coverslips were washed extensively and fixed on slides, and images were taken under an LSM800 confocal microscope (Carl Zeiss MicroImaging GmbH, Jena, Germany).
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