Anti Rabbit IgG, Goat, SPICA DyeTM 647 Conjugated
- for Immunochemistry
- Manufacturer :
- FUJIFILM Wako Pure Chemical Corporation
- Storage Condition :
- Keep at 2-10 degrees C.
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500uL
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Product Overview
Anti Rabbit IgG, Goat, SPICA Dye™ 647 Conjugated is a secondary antibody conjugated with SPICA Dye™, a novel ultra-photostable fluorescent dye developed by Fujifilm. Compared to conventional fluorescent dyes, SPICA Dye™ is significantly more resistant to photobleaching. This antibody is suitable for both immunohistochemistry and immunocytochemistry.
Antibody Information
| Clonality | Polyclonal |
|---|---|
| Antigen | Rabbit IgG |
| Host | Goat |
| Formulation | PBS, 0.05% Sodium azide |
| Conjugate | SPICA Dye™ 647 |
| Cross-reactivity | Rabbit |
| Application | Immunohistochemistry (frozen section) 1:500-1,000 Immunocytochemistry 1:500-1,000 |
Excitation/Emission
SPICA Dye™ 647
Protocol (Example)
Immunohistochemistry of Mouse Brain Sections using Anti-Iba1 Antibody and Anti Rabbit IgG, Goat, SPICA Dye™ 647 Conjugated
- 1.Perfusion fixation
- Perfuse with 0.1% sodium citrate/PBS to remove blood, followed by perfusion with 4% PFA.
- 2.Post-fixation
- 4% PFA (24 hours)
- 3.Immersion in sucrose
- 30% Sucrose/PBS (4 °C, from overnight to approximately 2 days)
- 4.Preparation of frozen sections
- After removing sucrose, prepare frozen blocks and cut 30 μm thick sections using a cryostat.
- 5.Blocking
- 5% BSA and 2% normal donkey serum in 0.3% Triton X-100/PBS (RT, 2 hours)
- 6.Primary antibody reaction
- Anti Iba1, Rabbit (for Immunocytochemistry) (Product Number 019-19741) 1:1,000 (4 °C, Overnight)
- 7.Washing
- 0.3% TritonX-100/PBS (5 minutes x 3)
- 8.Secondary antibody reaction
- Anti Rabbit IgG, Goat, SPICA Dye™ 647 Conjugated (This product) 1:500 in 0.3% Triton X-100/PBS (RT, 2 hours)
- 9.Washing
- 0.3% TritonX-100/PBS (5 minutes x 3)
- 10.Mounting
- The sample is sealed in mounting medium (ex. Invitrogen™ ProLong Glass) and stored at 4 °C in a dark place.
Data
Performance Data
Photostability Evaluation Using Xenon Light Resistance Test Compared to Conventional Dyes
Using a xenon light resistance tester equipped with a 500 W xenon lamp and a glass filter, PBS solutions containing goat anti-rabbit IgG antibodies conjugated with either SPICA Dye™ 647 or Alexa Fluor® 647 (Jackson ImmunoResearch Inc., Product Number 111-005-003) were continuously subjected to xenon light irradiation. Changes in absorbance at the maximum absorption wavelength were plotted to compare the photostability of the two dyes.
[Result]
The SPICA Dye™ 647-conjugated antibody demonstrated greater photostability than the antibody conjugated with the competitor’s fluorescent dye.
Photostability Evaluation by Widefield Microscopy
Photobleaching of near infrared fluorescent dyes for immunostaining Ki-67 in HeLa cells (widefield illumination).
(A) Representative images. Scale bar, 50 μm.
(B) Photobleaching curves.
(Alexa647_1-6); Goat anti-rabbit IgG (from Fuji*), Donkey anti-rabbit IgG (Jackson, 711-605-152), Goat anti-rabbit IgG (Thermo, A21244), Donkey anti-rabbit IgG (Thermo, A31573), Goat anti-rabit IgG (abcam, ab150083), Donkey anti-rabbit IgG (abcam, ab150075)
Microscope: IX83 P2ZF
Objective lens: UPLXAPO 40X/0.95 (NA=0.95) (Evident)
Ex/Em cube: Cy5-A-Basic (Semrock)
* Fuji: FUJIFILM Corp.
Photostability Evaluation by Confocal Microscopy
Photobleaching of near infrared fluorescent dyes for immunostaining Ki-67 in HeLa cells by confocal microscopy.
(A) Representative images.
Microscope: FV1200 CLSM
Objective lens: XLFLUOR 4X/340 (NA=0.28)
Illumination: 635 nm laser (0.81 W/cm2)
Scan Speed: 4.0 ms/pixel
Zoom: x 4.0
(B) Photobleaching curves.
Microscope: FV3000 CLSM (Evident)
Objective lens: UPLSAPO 40X(NA=0.95) (Evident)
Illumination: 640 nm laser (3.24 W/cm2)
Scan Speed: 2.0 ms/pixel
Zoom: x 2
* Alexa Fluor® is a registered trademark of Invitrogen Corporation.
FAQ
About antibody
- How is this antibody purified?
- This antibody is purified by antigen affinity chromatography.
- What is the concentration of the antibody?
- 1.4-1.6 mg/mL (as a protein concentration). The actual concentration is indicated on the label.
Overview / Applications
| Outline | Secondary antibodies are versatile and can be used in a variety of detection systems, including immunostaining. Anti-Rabbit IgG secondary antibodies are affinity-purified antibodies with characterized specificity for rabbit IgG and are useful in the detection or sorting of its target. Anti Rabbit IgG, Goat, SPICA DyeTM 647 Conjugated is conjugated to SPICA DyeTM 647 fluorescent dye under optimal conditions. Since this product is a polyclonal antibody and multiple secondary antibodies can bind to one primary antibody, signal amplification can also be expected. SPICA DyeTM 647 (Excitation:651 nm, Emission:670 nm) has superior light resistance compared to existing fluorescent dyes and is useful for experiments such as transparency and ultraresolution microscopy. [Class] Goat IgG [Species cross reactivity] Rabbit [Application] Immunocytochemistry 1:500 - 1:1,000 Immunohistochemistry (frozen section) 1:500 - 1:1,000 Optimal concentration should be determined by each laboratory for each application. |
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Property
| Appearance | Liquid |
|---|---|
| Concentration | Described on the label Protein : 1.4 - 1.6mg/mL |
Manufacturer Information
Alias
For research use or further manufacturing use only. Not for use in diagnostic procedures.
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