Ab-10 Rapid HiLyte Fluor 647 Labeling Kit

- Overview
  - Ab-10 Rapid HiLyte Fluor* 647 Labeling Kit enables rapid (in less than 30 min) and easy labeling of HiLyte Fluor 647 to 10 μg antibody. Reactive HiLyte Fluor 647 (a component of the kit) has succinimidyl ester group, that can easily make a covalent bond with an amino group of the target antibody without any activation process. This kit contains all the necessary reagents to prepare a fluorescein-labeled antibody.
    
    Please contact us at for specific academic pricing.
- Properties
  - Categories
    
    Fluorophore labeling
    
    Storage
    
    0-5°C
    
    Shipping
    
    ambient temperature
    
    * For research use only
- Applications
  - Application
    
    FCM; Microscopy
    
    Application Description
    
    Mitochondria Immunostaining
    1. HeLa cells were seeded on a μ-slide 8 well (ibidi) and cultured overnight at 37 °C in a 5% CO2 incubator.
    2. The cells were washed with PBS three times, and 4% paraformaldehyde in PBS was added to the μ-slide.
    3. The μ-slide was incubated at room temperature for 15 minutes.
    4. The cells were washed with PBS three times, and 1% Triton-X in PBS was added to the μ-slide.
    5. The μ-slide was incubated at room temperature for 30 minutes.
    6. Once the cells were washed with PBS three times, a blocking solution prepared with PBS was added to the μ-slide.
    7. The cells were then incubated at room temperature for 1 hour.
    8. Biotin conjugated anti-mitochondria antibody was diluted 50 times with the blocking solution. 　
    ※Anti-mitochondria antibody was purchased from Abcam (Product Code: ab3298) .
    9. The supernatant was discarded and the solution (step 8) was added to the μ-slide.
    10. The μ-slide was incubated at 0-5°C overnight.
    11. The supernatant was discarded and the cells were washed using PBS-T three times.
    12. 0.2 µg/ml peroxidase conjugated streptavidin was added to the μ-slide.
    13. The μ-slide was incubated at room temperature for 1 hour.
    14. The supernatant was discarded and the cells were washed using PBS-T three times.
    15. The cells were washed using Tris buffer (TB, 50 mmol/l, pH 7.5) three times.
    16. The supernatant was discarded and DAB solution [0.2 mg/ml DAB (SKU:D006), 0.003% H2O2 , 50 mmol/l Tris (pH 7.5)] was added to the μ-slide.
    17. The μ-slide was incubated at room temperature for 10 minutes.
    18. After the cells were washed using TB three times, TB was added to the μ-slide.
    19.The cells were observed under a microscope.

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Manual

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