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Four-color fluorescence in situ hybridization on a Drosophila embryo. A late blastoderm stage (nuclear cycle 14) embryo was probed with four different RNA probes. Blue: sog labeled with DNP, followed by a rabbit anti–dinitrophenyl-KLH IgG antibody (Cat. No. A6430) detected with an Alexa Fluor® 647 chicken anti–rabbit IgG antibody (Cat. No. A21443). Green: ind labeled with biotin, followed by streptavidin HRP and Alexa Fluor® 350 tyramide (TSA Kit ##27, Cat. No. T20937). Red: msh labeled with digoxigenin followed by sheep anti-digoxigenin antibody detected with an Alexa Fluor® 488 donkey anti–sheep IgG antibody (Cat. No. A11015). Yellow: sna labeled with fluorescein followed by mouse anti-fluorescein antibody detected with an Alexa Fluor® 555 goat anti–mouse IgG antibody (Cat. No. A21424). Image contributed by Dave Kosman and Ethan Bier, University of California, San Diego.
|Tested species reactivity||Mouse|
|Published species reactivity||Not Applicable|
|Host / Isotype||Goat / IgG|
|Immunogen||Gamma Immunoglobins Heavy and Light chains|
|Conjugate||Alexa Fluor® 555|
|Storage buffer||PBS, pH 7.5|
|Contains||5mM sodium azide|
|Storage Conditions||4° C, store in dark|
|Cross Adsorption||Against bovine IgG, goat IgG, rabbit IgG, rat IgG, human IgG and human serum|
|Antibody Form||Whole Antibody|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||1-10 µg/mL|
|Immunocytochemistry (ICC)||1-10 µg/mL|
|Immunofluorescence (IF)||1-10 µg/mL|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
To minimize cross-reactivity, these goat anti-mouse IgG (H+L) whole secondary antibodies have been affinity purified and cross-adsorbed against bovine IgG, goat IgG, rabbit IgG, rat IgG, human IgG, and human serum. Cross-adsorption or pre-adsorption is a purification step to increase specificity of the antibody resulting in higher sensitivity and less background staining. The secondary antibody solution is passed through a column matrix containing immobilized serum proteins from potentially cross-reactive species. Only the nonspecific-binding secondary antibodies are captured in the column, and the highly specific secondaries flow through. The benefits of this extra step are apparent in multiplexing/multicolor-staining experiments (e.g., flow cytometry) where there is potential cross-reactivity with other primary antibodies or in tissue/cell fluorescent staining experiments where there are may be the presence of endogenous immunoglobulins.
Alexa Fluor dyes are among the most trusted fluorescent dyes available today. Invitrogen™ Alexa Fluor 555 dye is a bright, orange-fluorescent dye with excitation ideally suited to the 555 nm laser line. For stable signal generation in imaging and flow cytometry, Alexa Fluor 555 dye is pH-insensitive over a wide molar range. Probes with high fluorescence quantum yield and high photostability allow detection of low-abundance biological structures with great sensitivity. Alexa Fluor 555 dye molecules can be attached to proteins at high molar ratios without significant self-quenching, enabling brighter conjugates and more sensitive detection. The degree of labeling for each conjugate is typically 2-8 fluorophore molecules per IgG molecule; the exact degree of labeling is indicated on the certificate of analysis for each product lot.
Using conjugate solutions: Centrifuge the protein conjugate solution briefly in a microcentrifuge before use; add only the supernatant to the experiment. This step will help eliminate any protein aggregates that may have formed during storage, thereby reducing nonspecific background staining. Because staining protocols vary with application, the appropriate dilution of antibody should be determined empirically. For the fluorophore-labeled antibodies a final concentration of 1-10 µg/mL should be satisfactory for most immunohistochemistry and flow cytometry applications.
We offer an extensive line of Invitrogen™ secondary antibody conjugates with well-characterized specificity and labeled with a wide selection of premium fluorescent dyes, including Invitrogen™ Alexa Fluor™ fluorescent dyes. Fluorescent secondary antibody conjugates are useful in the detection, sorting, or purification of its specified target and ideal for fluorescence microscopy and confocal laser scanning microscopy, flow cytometry, and fluorescent western detection. The breadth of fluorescent markers we offer allows our reagents to be tailored to almost any fluorescent detection system.
Secondary antibodies may be provided in three formats: whole IgG, divalent F(ab')2 fragments, and monovalent Fab fragments. Because of the high degree of conservation in the structure of many immunoglobulin domains, most class-specific secondary antibodies must be affinity-purified and cross-adsorbed to achieve minimal cross-reaction with other immunoglobulins.
Our secondary antibody conjugates are most commonly prepared by immunizing the host animal with a pooled population of immunoglobulins from the target species and can be further purified and modified (e.g., immunoaffinity chromatography, antibody fragmentation, label conjugation, etc.) to generate highly specific reagents. In the first round of purification, whole immunoglobulins binding to the immunizing antibody are recovered and mainly consist of the ~150-kDa IgG class. Further purification, for example, with Protein A or G, removes all unwanted immunoglobulin classes except the affinity-purified antibodies that react with the target-specific immunoglobulin heavy and/or light chains.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
Rivaroxaban and apixaban reduce hemorrhagic transformation after thrombolysis by protection of neurovascular unit in rat.
A-21424 was used in immunohistochemistry - frozen section to assess the effects of tissue-type plasminogen activator treatment after oral anticoagulation with rivaroxaban or apixaban compared with warfarin or placebo
|Kono S,Yamashita T,Deguchi K,Omote Y,Yunoki T,Sato K,Kurata T,Hishikawa N,Abe K||Stroke; a journal of cerebral circulation (45:2404)||2014|
|Not Applicable||Not Cited||The complexity of phosphorylated H2AX foci formation and DNA repair assembly at DNA double-strand breaks.||Nakamura AJ,Rao VA,Pommier Y,Bonner WM||Cell cycle (Georgetown, Tex.) (9:389)||2010|
|Not Applicable||Not Cited||High efficacy of a Listeria-based vaccine against metastatic breast cancer reveals a dual mode of action.||Kim SH,Castro F,Paterson Y,Gravekamp C||Cancer research (69:5860)||2009|
|Not Applicable||Not Cited||Simultaneous detection of mRNA and protein stem cell markers in live cells.||Rhee WJ,Bao G||BMC biotechnology (9:null)||2009|
|Not Applicable||Not Cited||Protective role of endogenous gangliosides for lysosomal pathology in a cellular model of synucleinopathies.||Wei J,Fujita M,Nakai M,Waragai M,Sekigawa A,Sugama S,Takenouchi T,Masliah E,Hashimoto M||The American journal of pathology (174:1891)||2009|
|Not Applicable||Not Cited||Early endosomes and endosomal coatomer are required for autophagy.||Razi M,Chan EY,Tooze SA||The Journal of cell biology (185:305)||2009|
|Not Applicable||Not Cited||Dual roles for an arginine-rich motif in specific genome recognition and localization of viral coat protein to RNA replication sites in flock house virus-infected cells.||Venter PA,Marshall D,Schneemann A||Journal of virology (83:2872)||2009|
|Not Applicable||Not Cited||High-efficiency labeling of sialylated glycoproteins on living cells.||Zeng Y,Ramya TN,Dirksen A,Dawson PE,Paulson JC||Nature methods (6:207)||2009|
|Not Applicable||Not Cited||A fluorimetry-based ssYFP secretion assay to monitor vasopressin-induced exocytosis in LLC-PK1 cells expressing aquaporin-2.||Nunes P,Hasler U,McKee M,Lu HA,Bouley R,Brown D||American journal of physiology. Cell physiology (295:C1476)||2008|
|Not Applicable||Not Cited||The transmembrane domain of the severe acute respiratory syndrome coronavirus ORF7b protein is necessary and sufficient for its retention in the Golgi complex.||Schaecher SR,Diamond MS,Pekosz A||Journal of virology (82:9477)||2008|
|Not Applicable||Not Cited||Subdiffraction multicolor imaging of the nuclear periphery with 3D structured illumination microscopy.||Schermelleh L,Carlton PM,Haase S,Shao L,Winoto L,Kner P,Burke B,Cardoso MC,Agard DA,Gustafsson MG,Leonhardt H,Sedat JW||Science (New York, N.Y.) (320:1332)||2008|
|Not Applicable||Not Cited||Rare steroid receptor-negative basal-like tumorigenic cells in luminal subtype human breast cancer xenografts.||Horwitz KB,Dye WW,Harrell JC,Kabos P,Sartorius CA||Proceedings of the National Academy of Sciences of the United States of America (105:5774)||2008|
|Not Applicable||Not Cited||Resolution of de novo HIV production and trafficking in immature dendritic cells.||Turville SG,Aravantinou M,Stössel H,Romani N,Robbiani M||Nature methods (5:75)||2008|
|Not Applicable||Not Cited||Comparison of hydroxylated print additives on antibody microarray performance.||Wu P,Grainger DW||Journal of proteome research (5:2956)||2006|