Sulfo-NHS-Biotin
(Synonyms: 生物素3-磺酸基-N-羟基琥珀酰亚胺钠盐,Sulfo-NHS Biotin) 目录号 : GC10804
Sulfo-NHS-Biotin是一种胺反应性生物素化试剂，主要用于对抗体、蛋白质和其他含伯胺(-NH2) 的大分子进行简单有效的生物素化标记。
Cas No.:119616-38-5
Sample solution is provided at 25 µL, 10mM.
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Quality Control & SDS
- View current batch:
- Purity: >95.00%
- COA (Certificate Of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
本方案仅提供一个指导，请根据您的具体需要进行修改。
1.溶液配制
（1）工作液：称量固体，用Milli-Q水或其他适合的缓冲液稀释到所需的工作浓度。
注意：固体避光保存在-20°C或-80°C。最佳的工作浓度请根据实际情况调整或参阅文献自行设置梯度浓度进行摸索。工作液必须现配现用。
2.采用Sulfo-NHS-Biotin进行细胞表面蛋白生物素化标记[1](来自文献,仅做参考)
（1）细胞刺激：HUVEC细胞在60 mm培养皿中培养，用LPS (1 μg/ml) 刺激细胞4小时。
（2）细胞表面蛋白的标记：将培养皿上的HUVEC单层与0.1 mg/ml Sulfo-NHS-Biotin的PBS溶液（补充有1.5 mM Ca2+）在 4°C下孵育30分钟，以最大程度地减少生物素化反应过程中细胞表面蛋白的内化。
（3）洗涤细胞：用冷的PBS溶液（补充有1.5 mM Ca2+）洗涤细胞，充分洗涤3次以除去过量的Sulfo-NHS-Biotin。
（4）继续培养细胞：用冷的DMEM溶液（补充有10% FBS）孵育细胞，以淬灭任何剩余的游离生物素试剂。
（5）洗涤细胞：细胞培养完成后经历另一次洗涤，以确保去除任何未结合的生物素。
（6）细胞裂解：细胞直接原位裂解，将细胞匀浆通过0.6 mm、24G针进行裂解，然后将裂解物以14000 rpm离心5分钟，以沉淀不溶性碎片。
（7）蛋白质分析：部分上清液作为总裂解液保存，使用NeutrAvidin琼脂糖分离生物素化蛋白质以进行后续分析。
注意：本实验提供了一项采用Sulfo-NHS-Biotin进行细胞表面蛋白生物素化标记的参考，Sulfo-NHS-Biotin的浓度和孵育时间需要根据细胞类型和所需的标记密度来优化。实验过程中尽量避免使用含有氨基酸、蛋白质或其他活性化合物的缓冲液，这些物质可能与生物素反应。
Sulfo-NHS-Biotin is an amine-reactive biotinylation reagent, mainly used for simple and effective biotinylation labeling of antibodies, proteins and other macromolecules containing primary amines (-NH2)[1]. Sulfo-NHS-Biotin is a simple NHS ester-activated biotinylation reagent composed of natural biotin valerate groups with an arm length of 13.5A[2]. Sulfo-NHS-Biotin is a water-soluble analog obtained by adding a negatively charged sulfonic acid group to the NHS-biotin structure[3]. Sulfo-NHS-Biotin can label surface proteins of the entire cell without penetrating the cell membrane[4].
References:
[1] Elia G. Biotinylation reagents for the study of cell surface proteins[J]. Proteomics, 2008, 8(19): 4012-4024.
[2]Cohen L, Walt D R. Evaluation of antibody biotinylation approaches for enhanced sensitivity of single molecule array (Simoa) immunoassays[J]. Bioconjugate Chemistry, 2018, 29(10): 3452-3458.
[3] Ahmed K K, Geary S M, Salem A K. Engineering the Surface of Cells Using Biotin–Avidin Chemistry[M]Micro-and Nanoengineering of the Cell Surface. William Andrew Publishing, 2014: 143-155.
[4]Reglero-Real N, Pérez-Gutiérrez L, Yoshimura A, et al. Autophagy modulates endothelial junctions to restrain neutrophil diapedesis during inflammation[J]. Immunity, 2021, 54(9): 1989-2004. e9.
Sulfo-NHS-Biotin是一种胺反应性生物素化试剂，主要用于对抗体、蛋白质和其他含伯胺(-NH2) 的大分子进行简单有效的生物素化标记[1]。Sulfo-NHS-Biotin是一种简单NHS酯活化的生物素化试剂，由天然生物素戊酸基团组成，臂长13.5A[2]。Sulfo-NHS-Biotin 是一种水溶性类似物，是通过在NHS-biotin结构上添加带负电荷的磺酸基而获得的[3]。Sulfo-NHS-Biotin可以对整个细胞的表面蛋白进行标记，不会渗透细胞膜[4]。
| Cas No. | 119616-38-5 | SDF | |
| 别名 | 生物素3-磺酸基-N-羟基琥珀酰亚胺钠盐,Sulfo-NHS Biotin | ||
| 化学名 | sodium;1-[5-[(3aS,6aR)-2-oxo-1,3,3a,4,6,6a-hexahydrothieno[3,4-d]imidazol-4-yl]pentanoyloxy]-2,5-dioxopyrrolidine-3-sulfonate | ||
| Canonical SMILES | C1C(C(=O)N(C1=O)OC(=O)CCCCC2C3C(CS2)NC(=O)N3)S(=O)(=O)[O-].[Na+] | ||
| 分子式 | C14H18N3NaO8S2 | 分子量 | 443.4 |
| 溶解度 | ≥ 22.17mg/mL in DMSO, ≥ 59.9 mg/mL in Water with ultrasonic | 储存条件 | Desiccate at -20°C, Unstable in solution, prepare and use immediately |
| General tips | 请根据产品在不同溶剂中的溶解度选择合适的溶剂配制储备液；一旦配成溶液，请分装保存，避免反复冻融造成的产品失效。 储备液的保存方式和期限：-80°C 储存时，请在 6 个月内使用，-20°C 储存时，请在 1 个月内使用。 为了提高溶解度，请将管子加热至37℃，然后在超声波浴中震荡一段时间。 |
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| Shipping Condition | 评估样品解决方案：配备蓝冰进行发货。所有其他可用尺寸：配备RT，或根据请求配备蓝冰。 | ||
| 制备储备液 | |||
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1 mg | 5 mg | 10 mg |
| 1 mM | 2.2553 mL | 11.2765 mL | 22.553 mL |
| 5 mM | 0.4511 mL | 2.2553 mL | 4.5106 mL |
| 10 mM | 0.2255 mL | 1.1276 mL | 2.2553 mL |
| 第一步：请输入基本实验信息（考虑到实验过程中的损耗，建议多配一只动物的药量） | ||||||||||
| 给药剂量 | mg/kg |  |
动物平均体重 | g | |
每只动物给药体积 | ul | |
动物数量 | 只 |
| 第二步：请输入动物体内配方组成（配方适用于不溶于水的药物；不同批次药物配方比例不同，请联系GLPBIO为您提供正确的澄清溶液配方） | ||||||||||
| % DMSO % % Tween 80 % saline | ||||||||||
| 计算重置 | ||||||||||
计算结果:
工作液浓度： mg/ml；
DMSO母液配制方法： mg 药物溶于 μL DMSO溶液（母液浓度 mg/mL，
体内配方配制方法：取 μL DMSO母液，加入 μL PEG300，混匀澄清后加入μL Tween 80，混匀澄清后加入 μL saline，混匀澄清。
1. 首先保证母液是澄清的；
2.
一定要按照顺序依次将溶剂加入，进行下一步操作之前必须保证上一步操作得到的是澄清的溶液，可采用涡旋、超声或水浴加热等物理方法助溶。
3. 以上所有助溶剂都可在 GlpBio 网站选购。
Cordycepin confers long-term neuroprotection via inhibiting neutrophil infiltration and neuroinflammation after traumatic brain injury
J Neuroinflammation 2021 Jun 15;18(1):137.34130727 PMC8207641
Background: The secondary injury caused by traumatic brain injury (TBI), especially white matter injury (WMI), is highly sensitive to neuroinflammation, which further leads to unfavored long-term outcomes. Although the cross-talk between the three active events, immune cell infiltration, BBB breakdown, and proinflammatory microglial/macrophage polarization, plays a role in the vicious cycle, its mechanisms are not fully understood. It has been reported that cordycepin, an extract from Cordyceps militaris, can inhibit TBI-induced neuroinflammation although the long-term effects of cordycepin remain unknown. Here, we report our investigation of cordycepin's long-term neuroprotective function and its underlying immunological mechanism. Methods: TBI mice model was established with a controlled cortical impact (CCI) method. Cordycepin was intraperitoneally administered twice daily for a week. Neurological outcomes were assessed by behavioral tests, including grid walking test, cylinder test, wire hang test, and rotarod test. Immunofluorescence staining, transmission electron microscopy, and electrophysiology recording were employed to assess histological and functional lesions. Quantitative-PCR and flow cytometry were used to detect neuroinflammation. The tracers of Sulfo-NHS-Biotin and Evans blue were assessed for the blood-brain barrier (BBB) leakage. Western blot and gelatin zymography were used to analyze protein activity or expression. Neutrophil depletion in vivo was performed via using Ly6G antibody intraperitoneal injection. Results: Cordycepin administration ameliorated long-term neurological deficits and reduced neuronal tissue loss in TBI mice. Meanwhile, the long-term integrity of white matter was also preserved, which was revealed in multiple dimensions, such as morphology, histology, ultrastructure, and electrical conductivity. Cordycepin administration inhibited microglia/macrophage pro-inflammatory polarization and promoted anti-inflammatory polarization after TBI. BBB breach was attenuated by cordycepin administration at 3 days after TBI. Cordycepin suppressed the activities of MMP-2 and MMP-9 and the neutrophil infiltration at 3 days after TBI. Moreover, neutrophil depletion provided a cordycepin-like effect, and cordycepin administration united with neutrophil depletion did not show a benefit of superposition. Conclusions: The long-term neuroprotective function of cordycepin via suppressing neutrophil infiltration after TBI, thereby preserving BBB integrity and changing microglia/macrophage polarization. These findings provide significant clinical potentials to improve the quality of life for TBI patients.
Assessment of solvent residues accessibility using three Sulfo-NHS-Biotin reagents in parallel: application to footprint changes of a methyltransferase upon binding its substrate
J Mass Spectrom 2008 Mar;43(3):360-70.17968972 10.1002/jms.1328
NHS-biotin modification as a specific lysine probe coupled to mass spectrometry detection is increasingly used over the past years for assessing amino acid accessibility of proteins or complexes as an alternative when well-established methods are challenged. We present a strategy based on usage in parallel of three commercially available reagents (Sulfo-NHS-Biotin, Sulfo-NHS-LC-biotin, and Sulfo-NHS-LC-LC-biotin) to efficiently assess the solvent accessibility of amino acids using MALDI-TOF mass spectrometry. The same qualitative pattern of reactivity was observed for these three reagents on the THUMPalpha protein at four reagent/polypeptide molar ratios (2 : 1, 6 : 1, 13 : 1, and 26 : 1). Peptide assignment of the detected ions gains in accuracy because of the triple redundancy due to specific increments of monoisotopic mass. These reagents are a good alternative to isotope labeling when using only a single MALDI-TOF mass spectrometer. We observed that hydroxyl groups of serine and tyrosine residues were also modified by these Sulfo-NHS-Biotin reagents. The low amount of protein required and the method's simplicity make this procedure accessible and affordable in order to obtain topological information on proteins difficult to purify. This method was used to identify two lysine residues of the TrmG10 methyltransferase from Pyrococcus abyssi that were differentially reactive, modified in the protein but not in the tRNA-protein complex.
The beta chain of the GPIIb molecule on ruminant leukocytes and platelets is not labelled by the Sulfo-NHS-Biotin method
Vox Sang 1995;69(3):248-9.8578739 10.1111/j.1423-0410.1995.tb02603.x
Glycoprotein (Gp)IIb/IIIa molecules have been immunoprecipitated from platelets and leukocytes of cattle, goat, horse, human, sheep and swine using specific monoclonal antibodies. The Sulfo-NHS-Biotin (sulfosuccinimidobiotin) method used to label the proteins has been found unsuitable for labelling the beta chain of the ruminant GPIIb/IIIa molecule. The beta chain was present on ruminant leukocytes and platelets when immunoprecipitates were silver stained.
Surface biotinylation of cytotoxic T lymphocytes for in vivo tracking of tumor immunotherapy in murine models
Cancer Immunol Immunother 2016 Dec;65(12):1545-1554.27722909 PMC5101144
Currently, there is no stable and flexible method to label and track cytotoxic T lymphocytes (CTLs) in vivo in CTL immunotherapy. We aimed to evaluate whether the sulfo-hydroxysuccinimide (NHS)-biotin-streptavidin (SA) platform could chemically modify the cell surface of CTLs for in vivo tracking. CD8+ T lymphocytes were labeled with Sulfo-NHS-Biotin under different conditions and then incubated with SA-Alexa647. Labeling efficiency was proportional to Sulfo-NHS-Biotin concentration. CD8+ T lymphocytes could be labeled with higher efficiency with Sulfo-NHS-Biotin in DPBS than in RPMI (P < 0.05). Incubation temperature was not a key factor. CTLs maintained sufficient labeling for at least 72 h (P < 0.05), without altering cell viability. After co-culturing labeled CTLs with mouse glioma stem cells (GSCs) engineered to present biotin on their surface, targeting CTLs could specifically target biotin-presenting GSCs and inhibited cell proliferation (P < 0.01) and tumor spheres formation. In a biotin-presenting GSC brain tumor model, targeting CTLs could be detected in biotin-presenting gliomas in mouse brains but not in the non-tumor-bearing contralateral hemispheres (P < 0.05). In vivo fluorescent molecular tomography imaging in a subcutaneous U87 mouse model confirmed that targeting CTLs homed in on the biotin-presenting U87 tumors but not the control U87 tumors. PET imaging with 89Zr-deferoxamine-biotin and SA showed a rapid clearance of the PET signal over 24 h in the control tumor, while only minimally decreased in the targeted tumor. Thus, sulfo-NHS-biotin-SA labeling is an efficient method to noninvasively track the migration of adoptive transferred CTLs and does not alter CTL viability or interfere with CTL-mediated cytotoxic activity.
Biotin as a probe of the surface of Ascaris suum developmental stages
Mol Biochem Parasitol 1990 Jun;41(1):45-52.2385267 10.1016/0166-6851(90)90095-4
Sulfo-NHS-Biotin (aqueous soluble) and NHS-biotin (organic soluble) labeled similar SDS-2ME (sodium dodecyl sulfate/beta-mercaptoethanol) soluble cuticular proteins of second stage larvae (L2) and third stage Ascaris suum larvae (L3). Comparable analysis of biotin-labeled fourth stage larvae (L4), young adults, and mature adult Ascaris suum revealed strong labeling of several SDS-2ME soluble cuticular proteins with NHS-biotin, while Sulfo-NHS-Biotin appeared to strongly label a single SDS-2ME soluble cuticular protein. Both biotin probes labeled only cuticular proteins, since no evidence of internal labeling was observed in any developmental stage examined by either electroblot analysis or by electron microscopy. Our data suggest a greater cuticular permeability to the organic soluble biotin reagent in the later developmental stages (greater than L3) of A. suum than to the aqueous soluble biotin reagent, and may indicate the presence of a hydrophobic barrier in the cuticle of the later stages of the parasite.