Dylight 647-Goat Anti-Rabbit IgG (H+L), SA-64702
Product Introduction
This product is a goat anti-Rabbit IgG (H+L) secondary antibody conjugated to Dylight 647, a bright and highly photostable far-red fluorescent dye. Secondary antibodies are essential reagents in immunofluorescence (IF), flow cytometry (FC), and other immunodetection techniques, providing specific detection of primary antibodies bound to target antigens. The antibody recognizes both the heavy and light chains of Rabbit IgG (H+L), ensuring broad reactivity with Rabbit IgG primary antibodies. Dylight 647 is a next-generation far-red fluorophore with excitation maximum at 654 nm and emission maximum at 674 nm, offering superior brightness, exceptional photostability, and minimal pH sensitivity compared to traditional far-red dyes such as Cy5. Its spectral properties are well-separated from common green and orange fluorophores (FITC, PE), making it an ideal choice for multi-color imaging and flow cytometry panels. The conjugate is supplied in a glycerol-containing storage buffer with BSA as a stabilizer and ProClin 300 as a preservative, ensuring long-term stability at -20¡ãC.
Product Features
1. Bright Far-Red Fluorophore: Dylight 647 offers superior fluorescence brightness and photostability, enabling sensitive detection of low-abundance targets.
2. Broad Rabbit IgG Reactivity: Recognizes both heavy and light chains (H+L) of Rabbit IgG, providing reactivity with all Rabbit IgG subclasses and IgM.
3. Multi-Color Compatible: Far-red emission (Emax=674 nm) is well-separated from blue, green, and red fluorophores, ideal for multi-color IF and FC panels.
4. Optimized for IF and FC: Validated for immunofluorescence and flow cytometry at dilutions of 1:200 to 1:1,000.
5. Long-Term Storage Stability: Formulated in glycerol-containing buffer with BSA stabilizer and ProClin 300 preservative for stability at -20¡ãC.
Specifications
Size: Available in 100 µL and 500 µL
Host Species: Goat
Target Species: Rabbit
Antibody Specificity: IgG (H+L), recognizes heavy and light chains
Conjugate: Dylight 647
Excitation/Emission: 654 nm / 674 nm
Recommended Dilution: 1:200 to 1:1,000 for IF and FC
Storage Buffer: PBS with 50% glycerol, 10 mg/mL BSA, 0.1% ProClin 300, pH 7.4
Storage and Stability
Storage Conditions: Store at -20¡ãC, protected from light. Stable for one year after shipment.
Shelf Life: The product is stable for 12 months from the date of shipment when stored as directed. Avoid repeated freeze-thaw cycles; for frequent use, aliquot and store at -20¡ãC.
Protocol (For Reference Only)
Important: Protect the fluorescent conjugate from light throughout handling and staining procedures. Centrifuge the vial briefly before opening to collect contents at the bottom.
1. Antibody Dilution: Dilute Dylight 647-Goat Anti-Rabbit IgG (H+L) in appropriate blocking buffer or antibody dilution buffer to a working concentration of 1:200 to 1:1,000 (e.g., 1 µL of conjugate in 200-1,000 µL of buffer). The optimal dilution should be determined empirically for each application and primary antibody.
2. Immunofluorescence (IF) Staining:
a. Following fixation, permeabilization, and blocking of cells or tissue sections, incubate with Rabbit primary antibody at the optimized concentration.
b. After primary antibody incubation and washing, add diluted Dylight 647 conjugate and incubate for 1 hour at room temperature in the dark.
c. Wash 3-5 times with PBS or TBST to remove unbound conjugate.
d. Counterstain with DAPI or Hoechst 33342 if desired, mount with anti-fade mounting medium, and image using a far-red filter set (excitation 630-650 nm, emission 660-680 nm).
3. Flow Cytometry (FC) Staining:
a. Following blocking of Fc receptors if required, incubate cells with Rabbit primary antibody at the optimized concentration.
b. After primary antibody incubation and washing, add diluted Dylight 647 conjugate and incubate for 30-60 minutes at 4¡ãC or room temperature in the dark.
c. Wash cells 2-3 times with flow cytometry buffer (PBS with 1% BSA or 2% FBS).
d. Resuspend cells in appropriate buffer and analyze by flow cytometry using the APC or Cy5 channel (excitation 633-640 nm laser, emission 660/20 nm filter).
4. Multi-Color Panel Design: When using Dylight 647 in multi-color experiments, ensure appropriate single-stain controls are included for compensation. Dylight 647 fluorescence is detected in the APC/Cy5 channel and is well-separated from FITC (green), PE (orange), and DAPI/Hoechst (blue) channels.
Precautions
1. Protect the fluorescent conjugate from light at all times during handling, staining, and storage. Prolonged light exposure causes photobleaching.
2. The recommended dilution range of 1:200 to 1:1,000 is a guideline; optimal dilution must be determined empirically for each primary antibody, cell type, and application.
3. The storage buffer contains 50% glycerol, which prevents freezing at -20¡ãC. The conjugate remains in liquid form; do not freeze-thaw by warming above 0¡ãC repeatedly.
4. If using for super-resolution microscopy or single-molecule imaging, pre-centrifuge the diluted conjugate at 10,000 x g for 5 minutes to remove potential aggregates.
5. For research use only. Not for use in diagnostic or therapeutic procedures.
FAQ (Simplified)
Q1: What is the recommended starting dilution for immunofluorescence?
A1: The recommended starting dilution is 1:200 to 1:500 for immunofluorescence. For abundant targets or bright primary antibodies, 1:1,000 may be sufficient. For low-abundance targets, a 1:200 dilution provides maximum sensitivity. Titration using your specific primary antibody and sample is recommended.
Q2: Can this antibody be used for Western blot?
A2: This conjugate is optimized for IF and FC. While it can be used for fluorescent Western blotting, signal detection may be less sensitive than with near-infrared dyes (e.g., IRDye 680) or HRP-based chemiluminescence, which are generally preferred for this application.
Q3: What are the advantages of Dylight 647 over Cy5?
A3: Dylight 647 offers comparable spectral properties to Cy5 but with superior brightness, greater photostability, and reduced pH sensitivity. This makes it particularly suitable for quantitative imaging and applications requiring extended illumination or imaging times.
Q4: How should I store this product to maintain stability?
A4: Store at -20¡ãC protected from light. The glycerol-containing buffer prevents freezing, so the conjugate remains liquid. The product is stable for one year after shipment. For frequent use, aliquot into smaller volumes to minimize repeated warming and light exposure.
Disclaimer
1. For Research Use Only. Not for use in diagnostic or therapeutic procedures.
2. Due to the variable nature of biological research, optimization of antibody dilution and staining conditions is recommended for specific primary antibodies and experimental systems.
3. This warranty is limited to the replacement of the product. The manufacturer assumes no liability for incidental or consequential damages, including loss of samples or data.
4. Wear appropriate protective clothing and gloves when handling this product.
Ordering Information
Catalog Number: SA-64702
Product Name: Dylight 647-Goat Anti-Rabbit IgG (H+L)
Size: 100 µL / 500 µL
Price:
100 µL: CNY ¥650.00 / USD $65.00 / EUR €78.00 / JPY ¥11700.00
500 µL: CNY ¥2580.00 / USD $258.00 / EUR €309.60 / JPY ¥46440.00
