ICG-Sulfo-OSu

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Telephone	1-800-990-8053
Fax	1-800-609-2943
Email	sales@aatbio.com
International	See distributors
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Shipping	Standard overnight for United States, inquire for international

Physical properties

Molecular weight	930.07
Solvent	DMSO

Spectral properties

Correction Factor (280 nm)	0.076
Extinction coefficient (cm ^-1 M ^-1)	230000
Excitation (nm)	789
Emission (nm)	813

Storage, safety and handling

H-phrase	H303, H313, H333
Hazard symbol	XN
Intended use	Research Use Only (RUO)
R-phrase	R20, R21, R22
Storage	Freeze (< -15 °C); Minimize light exposure
UNSPSC	12171501

Overview SDS Protocol

Example protocol

AT A GLANCE

Molecular Weight: 930.07
Solvent: DMSO
Extinction Coefficient: 230,000 cm-1M-1
Excitation/Emission: 780/800 nm
CF at 260/280 nm: 0.113/0.073
Important Extinction coefficient are at their maximum absorption wavelength. CF at 260 nm is the correction factor used for eliminating the dye contribution to the absorbance at 260 nm (for oligo and nucleic acid labeling). CF at 280 nm is the correction factor used for eliminating the dye contribution to the absorbance at 280 nm (for peptide and protein labeling). Fluorescence intensity is significantly increased upon coupled to proteins. This labeling protocol was developed for the conjugate of Goat anti-mouse IgG with ICG. You might need further optimization for your particular proteins.

PREPARATION OF STOCK SOLUTIONS

Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles.

1. Protein Stock Solution (Solution A)

Mix 100 µL of a reaction buffer (e.g., 1 M sodium carbonate solution or 1 M phosphate buffer with pH ~9.0) with 900 µL of the target protein solution (e.g. antibody, protein concentration ≥2 mg/ml if possible) to give 1 mL protein labeling stock solution. The pH of the protein solution should be 8.5 ± 0.5. If the pH of the protein solution is lower than 8.0, adjust the pH to the range of 8.0 - 9.0 using 1 M sodium bicarbonate solution or 1 M pH 9.0 phosphate buffer. The protein should be dissolved in 1X phosphate buffered saline (PBS), pH 7.2 - 7.4. If the protein is dissolved in Tris or glycine buffer, it must be dialyzed against 1X PBS, pH 7.2 - 7.4, to remove free amines or ammonium salts (such as ammonium sulfate and ammonium acetate) that are widely used for protein precipitation. Impure antibodies or antibodies stabilized with bovine serum albumin (BSA) or gelatin will not be labeled well. The presence of sodium azide or thimerosal might also interfere with the conjugation reaction. Sodium azide or thimerosal can be removed by dialysis or spin column for optimal labeling results. For optimal labeling efficiency, the final protein concentration range of 2 - 10 mg/mL is recommended, with the conjugation efficiency significantly reduced if less than 2 mg/mL.

2. Dye Stock Solution (Solution B)

Add anhydrous DMSO into the vial of ICG dyes to make a 10 - 20 mM stock solution. Mix well by pipetting or vortex. Prepare the dye stock solution before starting the conjugation. Use promptly. Extended storage of the dye stock solution may reduce dye activity.

SAMPLE EXPERIMENTAL PROTOCOL

Determine the optimal dye/protein ratio (optional):

Use 10:1 molar ratio of Solution B (dye)/Solution A (protein) as the starting point: Add 5 µl of the dye stock solution (Solution B, assuming the dye stock solution is 10 mM) into the vial of the protein solution (95 µl of Solution A) with effective shaking. The concentration of the protein is ~0.05 mM assuming the protein concentration is 10 mg/mL and the molecular weight of the protein is ~200KD. The concentration of the DMSO in the protein solution should be <10%.
Run conjugation reaction.
Repeat Step 2 with the molar ratios of Solution B/Solution A at 5:1; 15:1 and 20:1 respectively.
Purify the desired conjugates using premade spin columns.
Calculate the dye/protein ratio (DOS) for the above 4 conjugates (if step 3 is done).
Run your functional tests of the above 4 conjugates to determine the best dye/protein ratio to scale up your labeling reaction.

Run conjugation reaction:

Add the appropriate amount of dye stock solution (Solution B) into the vial of the protein solution (Solution A) with effective shaking. The best molar ratio of Solution B/Solution is determined above. If skipped, we recommend using 10:1 molar ratio of Solution B (dye)/Solution A (protein).
Continue to rotate or shake the reaction mixture at room temperature for 30-60 minutes.

Purify the conjugation:The following protocol is an example of dye-protein conjugate purification by using a Sephadex G-25 column.

Prepare Sephadex G-25 column according to the manufacture instruction.
Load the reaction mixture to the top of the Sephadex G-25 column.
Add PBS (pH 7.2 - 7.4) as soon as the sample runs just below the top resin surface.
Add more PBS (pH 7.2 - 7.4) to the desired sample to complete the column purification. Combine the fractions that contain the desired dye-protein conjugate. For immediate use, the dye-protein conjugate need be diluted with staining buffer, and aliquoted for multiple uses. For longer term storage, dye-protein conjugate solution need be concentrated or freeze dried.

Calculators

Common stock solution preparation

Table 1. Volume of DMSO needed to reconstitute specific mass of ICG-Sulfo-OSu to given concentration. Note that volume is only for preparing stock solution. Refer to sample experimental protocol for appropriate experimental/physiological buffers.

	0.1 mg	0.5 mg	1 mg	5 mg	10 mg
1 mM	107.519 µL	537.594 µL	1.075 mL	5.376 mL	10.752 mL
5 mM	21.504 µL	107.519 µL	215.038 µL	1.075 mL	2.15 mL
10 mM	10.752 µL	53.759 µL	107.519 µL	537.594 µL	1.075 mL

Molarity calculator

Enter any two values (mass, volume, concentration) to calculate the third.

Mass (Calculate)		Molecular weight		Volume (Calculate)		Concentration (Calculate)		Moles
	/		=		x		=

Spectrum

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Spectral properties

Correction Factor (280 nm)	0.076
Extinction coefficient (cm ^-1 M ^-1)	230000
Excitation (nm)	789
Emission (nm)	813

Product Family

Name	Excitation (nm)	Emission (nm)	Extinction coefficient (cm ^-1 M ^-1)	Correction Factor (280 nm)	Correction Factor (260 nm)
ICG-PEG12-OSu	789	813	230000	0.076	-
ICG Xtra-OSu	790	814	230000	0.055	0.09

Images

Figure 1. Chemical structure for ICG-Sulfo-OSu

Citations

View all 17 citations: Citation Explorer

Targeted delivery of cathepsin-activatable near-infrared fluorescence probe for ultrahigh specific imaging of peritoneal metastasis
Authors: Lu, Jialiang and Guo, Yu and Hao, Huimin and Ma, Junjie and Lu, Yang and Sun, Yue and Shi, Zheng and Dong, Xiaowu and Zhang, Bo and Fang, Luo and others,
Journal: European Journal of Medicinal Chemistry (2023): 115909

pH-gated nanoparticles selectively regulate lysosomal function of tumour-associated macrophages for cancer immunotherapy
Authors: Tang, Mingmei and Chen, Binlong and Xia, Heming and Pan, Meijie and Zhao, Ruiyang and Zhou, Jiayi and Yin, Qingqing and Wan, Fangjie and Yan, Yue and Fu, Chuanxun and others,
Journal: Nature Communications (2023): 5888

Comparison of Near-Infrared Imaging Agents Targeting the PTPmu Tumor Biomarker
Authors: Johansen, Mette L and Vincent, Jason and Rose, Marissa and Sloan, Andrew E and Brady-Kalnay, Susann M
Journal: Molecular Imaging and Biology (2023): 1--14

Quantitative imaging of intracellular nanoparticle exposure enables prediction of nanotherapeutic efficacy
Authors: Yin, Qingqing and Pan, Anni and Chen, Binlong and Wang, Zenghui and Tang, Mingmei and Yan, Yue and Wang, Yaoqi and Xia, Heming and Chen, Wei and Du, Hongliang and others,
Journal: Nature communications (2021): 1--13

Detection of Lymph Node Metastases by Ultra-pH-Sensitive Polymeric Nanoparticles
Authors: Bennett, Zachary T and Feng, Qiang and Bishop, Justin A and Huang, Gang and Sumer, Baran D and Gao, Jinming
Journal: Theranostics (2020): 3340

Detection of lymph node metastases by ultra-pH-sensitive polymeric nanoparticles
Authors: Bennett, Zachary T and Feng, Qiang and Bishop, Justin A and Huang, Gang and Sumer, Baran D and Gao, Jinming
Journal: Theranostics (2020): 3340

Ultrasmall hybrid protein--copper sulfide nanoparticles for targeted photoacoustic imaging of orthotopic hepatocellular carcinoma with a high signal-to-noise ratio
Authors: Yan, Huixiang and Chen, Jingqin and Li, Ying and Bai, Yuanyuan and Wu, Yunzhu and Sheng, Zonghai and Song, Liang and Liu, Chengbo and Zhang, Hai
Journal: Biomaterials science (2019): 92--103

Tracking and Imaging of Transplanted Stem Cells in Animals
Authors: Chikate, Tanmayee Rajeev and Tang, Liping
Journal: (2019)

Autophagy promotes hepatic differentiation of hepatic progenitor cells by regulating the Wnt/β-catenin signaling pathway
Authors: Ma, Zhenzeng and Li, Fei and Chen, Liuying and Gu, Tianyi and Zhang, Qidi and Qu, Ying and Xu, Mingyi and Cai, Xiaobo and Lu, Lungen
Journal: Journal of molecular histology (2019): 1--16

Hybrid MoSe 2--indocyanine green nanosheets as a highly efficient phototheranostic agent for photoacoustic imaging guided photothermal cancer therapy
Authors: Chen, Jingqin and Li, Xueshen and Liu, Xiaoyang and Yan, Huixiang and Xie, Zhihua and Sheng, Zonghai and Gong, Xiaojing and Wang, Lidai and Liu, Xin and Zhang, Peng and others,
Journal: Biomaterials science (2018): 1503--1516

References

View all 193 references: Citation Explorer

Ultrastaging of colon cancer by sentinel node biopsy using fluorescence navigation with indocyanine green
Authors: Hirche C, Mohr Z, Kneif S, Doniga S, Murawa D, Strik M, Hunerbein M.
Journal: Int J Colorectal Dis (2012): 319

Sentinel lymph node biopsy using real-time fluorescence navigation with indocyanine green in cutaneous head and neck/lip mucosa melanomas
Authors: Hayashi T, Furukawa H, Oyama A, Funayama E, Saito A, Yamao T, Yamamoto Y.
Journal: Head Neck (2012): 758

Efficacy of indocyanine green videography and real-time evaluation by FLOW 800 in the resection of a spinal cord hemangioblastoma in a child
Authors: Ueba T, Abe H, Matsumoto J, Higashi T, Inoue T.
Journal: J Neurosurg Pediatr (2012): 428

Concentration of indocyanine green does not significantly influence lymphatic function as assessed by near-infrared imaging
Authors: Aldrich MB, Davies-Venn C, Angermiller B, Robinson H, Chan W, Kwon S, Sevick-Muraca EM.
Journal: Lymphat Res Biol (2012): 20

Comparative Study of the Optical and Heat Generation Properties of IR820 and Indocyanine Green
Authors: Fern, undefined and ez-Fern, undefined and ez A, Manch and a R, Lei T, Carvajal DA, Tang Y, Zahid Raza Kazmi S, McGoron AJ.
Journal: Mol Imaging (2012): 99

Sentinel lymph node biopsy using a new indocyanine green fluorescence imaging system with a colour charged couple device camera for oral cancer
Authors: Iwai T, Maegawa J, Hirota M, Tohnai I.
Journal: Br J Oral Maxillofac Surg. (2012)

Usefulness of indocyanine green angiography to depict the distant retinal vascular anomalies associated with branch retinal vein occlusion causing serous macular detachment
Authors: Ueda T, Gomi F, Suzuki M, Sakaguchi H, Sawa M, Kamei M, Nishida K.
Journal: Retina (2012): 308

Application of indocyanine green videoangiography in surgery for spinal vascular malformations
Authors: Misra BK, Pur and are HR., undefined
Journal: J Clin Neurosci. (2012)

Indocyanine-Green Angiography Findings in Susac's Syndrome
Authors: Balaskas K, Guex-Crosier Y, Borruat FX.
Journal: Klin Monbl Augenheilkd (2012): 426

Gold nanomaterials conjugated with indocyanine green for dual-modality photodynamic and photothermal therapy
Authors: Kuo WS, Chang YT, Cho KC, Chiu KC, Lien CH, Yeh CS, Chen SJ.
Journal: Biomaterials (2012): 3270