| Product Specific InformationAlexa Fluor dyes are among the most trusted fluorescent dyes available today. Invitrogen™ Alexa Fluor™ 488 dye is a bright, green-fluorescent dye with excitation ideally suited to the 488 nm laser line. For stable signal generation in imaging and flow cytometry, Alexa Fluor 488 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 488 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.The goat anti-rabbit IgG whole 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 with Protein A or G removes all immunoglobulin classes except IgG such that the affinity-purified antibodies react with IgG heavy chains and all classes of immunoglobulin light chains from rabbit. To minimize cross-reactivity, these goat anti-rabbit whole antibodies have been cross-adsorbed against human IgG, human serum, mouse IgG, mouse serum, and bovine 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 where there is potential cross-reactivity with other primary antibodies or in immunohistochemistry experiments where there are may be the presence of endogenous immunoglobulins. For a highly cross-adsorbed secondary antibody equivalent (or equivalent secondary antibody preparation), please see product catalog number: A11034.Using conjugate solutions: Centrifuge the protein conjugate solution briefly in a microcentrifuge before use; add only the supernatant to the experiment. This step will 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.Background/Target InformationWe 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.For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization. |
| 数据 |
Rabbit IgG (H+L) Cross-Adsorbed Secondary Antibody (A-11008) in IFMitochondria were stained with the MitoTracker® Red CMXRos reagent (Product # M-7512). Peroxisomal labeling was achieved with a primary antibody directed against PMP70, visualized using green-fluorescent Alexa Fluor® 488 goat anti-rabbit IgG (Product # A-11008). The nucleus was stained with blue-fluorescent DAPI (Product # D1306, D3571, D21490). Rabbit IgG (H+L) Cross-Adsorbed Secondary Antibody (A-11008) in IFOCT-embedded sections were incubated with a rabbit primary antibody to PGC7 (a germ cell marker) and subsequently visualized using an Alexa Fluor® 488 goat anti-rabbit IgG antibody (Product # A-11008). The image was contributed by Masatake Sato, Osaka University. Rabbit IgG (H+L) Cross-Adsorbed Secondary Antibody (A-11008) in IFPseudocolored green-fluorescent labeling represents a fluorescein-labeled cRNA probe detected using a rabbit anti-fluorescein/Oregon Green® primary antibody (Product # A-889) and an Alexa Fluor® 488 dye-labeled anti-rabbit secondary antibody (Product # A-11008). Pseudocolored yellow- and red-fluorescent labeling represents a biotinylated cRNA probe detected using HRP-streptavidin and Alexa Fluor® 568 tyramide (TSA Kit #24, Product # T-20934). Pseudocolored blue-fluorescent labeling represents a digoxigenin-labeled cRNA probe detected using a mouse anti-digoxigenin primary antibody in conjunction with an Alexa Fluor® 647 dye-labeled anti-mouse secondary antibody (Product # A-21235). The image was contributed by Ethan Bier and David Kosman, University of California, San Diego. Rabbit IgG (H+L) Cross-Adsorbed Secondary Antibody (A-11008) in IFImmunofluorescence analysis of Goat anti-Rabbit IgG (H+L) Cross-Adsorbed Secondary Antibody Alexa Fluor® 488 conjugate was performed using HeLa cells stained with alpha Tubulin Rabbit Polyclonal Antibody (Product # PA5-16891). The cells were fixed with 4% paraformaldehyde for 10 minutes, permeabilized with 0.1% Triton™ X-100 for 10 minutes, blocked with 1% BSA for 1 hour and labeled with 2 µg/ml Rabbit primary antibody for 3 hours at room temperature. Goat anti-Rabbit IgG (H+L) Cross-Adsorbed Secondary Antibody Alexa Fluor® 488 conjugate (Product # A-11008) was used at a concentration of 4 µg/ml in phosphate buffered saline containing 0.2% BSA for 45 minutes at room temperature, for detection of alpha Tubulin in the cytoplasm (Panel a: green). Nuclei (Panel b: blue) were stained with DAPI in SlowFade® Gold Antifade Mountant (Product # S36938). F-actin was stained with Rhodamine Phalloidin (Product # R415, 1:300) (Panel c: red). Panel d represents the composite image. No nonspecific staining was observed with the secondary antibody alone (panel f), or with an isotype control (panel e). The images were captured at 60X magnification. |
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| 1.Skeletal muscleFailed reinnervation in aging skeletal muscle."A-11008 was used in immunohistochemistry - frozen section to discuss the age-related presence of denervated myofibers and accelerated muscle atrophy"Authors Aare S,Spendiff S,Vuda M,Elkrief D,Perez A,Wu Q,Mayaki D,Hussain SN,Hettwer S,Hepple RT2.Molecular vision mice."A11008 was used in immunohistochemistry - frozen section to evaluate the mechanisms of retinal degeneration"Authors Dong E,Bachleda A,Xiong Y,Osawa S,Weiss ER3.Human molecular geneticsIncreased mitophagy in the skeletal muscle of spinal and bulbar muscular atrophy patients."A-11008 was used in immunohistochemistry - frozen section to characterize muscle biopsy specimens derived from patients with spinal and bulbar muscular atrophy"Authors Borgia D,Malena A,Spinazzi M,Desbats MA,Salviati L,Russell AP,Miotto G,Tosatto L,Pegoraro E,Sorarù G,Pennuto M,Vergani L4.PloS oneEffects of a skin-massaging device on the ex-vivo expression of human dermis proteins and in-vivo facial wrinkles."A11008 was used in immunohistochemistry - frozen section to characterize the effects of mild skin massage on cell behavior"Authors Caberlotto E,Ruiz L,Miller Z,Poletti M,Tadlock L5.Frontiers in cellular neuroscience"A-11008 was used in immunohistochemistry to evaluate commercially available adeno-associated virus serotypes for their ability to transduce primary rat astrocytes"Authors Schober AL,Gagarkin DA,Chen Y,Gao G,Jacobson L,Mongin AA |
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