Study on the Properties and Applications of Biotin-SS-NHS Ester
1. Basic Information and Structural Features of the Compound
Biotin-SS-NHS Ester (CAS No: 122266-55-1) is a biochemical reagent with a unique molecular structure. The molecular formula is C19H28N4O6S3, with a molecular weight of 504.64, and it typically has a purity standard above 95%. It appears as a white to off-white solid powder. Its molecular structure consists of three key functional units: biotin group, disulfide bond linker, and N-hydroxysuccinimide ester active end.
As a member of the vitamin B family, biotin serves as an essential cofactor for key metabolic enzymes such as carboxylases due to its high affinity binding characteristic with streptavidin (Kd≈10^-15M), making it the gold standard in biomarker fields. The disulfide bond (-SS-) acts as a dynamic covalent bond within the molecule that provides chemical stability while allowing cleavage under reducing conditions; this feature is significant for controlled release applications. The NHS ester end group is a classic reactive amino reaction site that can efficiently condense with primary amines in proteins or peptides under neutral to weakly alkaline conditions (pH 7.2–8.5), forming stable amide bonds.
2. Physicochemical Properties and Stability Parameters
This compound exhibits several special physicochemical properties. Its water solubility significantly surpasses traditional biotin reagents primarily due to hydrophilicity brought by sulfonate groups in its structure. Experimental data show that at 25°C, its solubility in water can reach up to 5–10 mg/mL; this property allows labeling reactions without using organic cosolvents like DMSO or DMF, greatly expanding its application range in sensitive biological systems.
Stability studies indicate that solid-state reagents maintain activity stability for over twelve months when stored away from light at -20°C in dry conditions. Notably, disulfide bonds may break down in reducing environments (e.g., presence of 1–10 mM DTT or TCEP); this characteristic is often designed into controlled release experimental systems. The half-life state solution closely relates to pH levels; under phosphate buffer saline at pH 7.4 at four degrees Celsius, activity can be maintained for four to six hours—hence fresh preparation before use is recommended.
3.Core Application Areas and Technical Advantages
3.1 Application in Proteomics Research In proteomics research field, this reagent has become an important labeling tool through covalently attaching biotin groups onto target proteins via NHS esters reacting with protein N-terminal or lysine residue ε-amino groups leading to efficient enrichment via streptavidin magnetic beads combined with mass spectrometry analysis techniques reaching detection limits down to fmol level compared against traditional biotination methods where presence of disulfides allows selective elution using reductants (like β-mercaptoethanol) avoiding denaturing conditions damaging protein structures during subsequent experiments. Particularly noteworthy are its unique values found within phosphoproteomic studies since biotin labeling does not interfere around phosphorylation sites while labeled phosphorylated peptide segments exhibit characteristic fragment peaks during mass spectrometric analyses rendering it ideal choice towards enriching & identifying phosphoproteins showing efficiency improvements between thirty-to-fifty percent based upon experimental data utilizing said strategies . 3..2 Innovative Applications In Immunoassay Technologies in immunoassays realm ,this reagent revolutionizes conventional antibody tagging strategies whereby optimizing label condition(typically employing sodium bicarbonate buffered system set @ ph8 .3 ;reaction time spanning two-four hours )allows introduction up three-five individual labels per each antibody molecule ensuring moderate density maintains sensitivity whilst preventing loss associated arising from excessive tagging causing decline overall functionality observed directly correlated against direct fluorescent markers thereby achieving signal amplification elevating detection sensitivity by factors ranging between two-three orders magnitude increase over prior methodologies employed across various platforms including flow cytometry showcasing distinctive advantages realized through reversible marking schemes enabling sequential rounds testing samples retaining integrity enhancing usage precious clinical specimens .
Four Operating Norms And Precautions
laboratory protocols must strictly adhere following guidelines :solubilizing utilize pre-chilled anhydrous DMSO preparing stock solutions concentrations ten-twenty millimolar subsequently aliquoted stored minus eighty degree celsius long-term preservation working dilutions suggested made utilizing non-amino buffers(pbs ,hepes )diluted final concentration zero point one-one millimolar marking reactions usually performed temperature ranges varying four twenty five degrees celcius duration lasting thirty minutes upto two hours post-reaction necessitating desalting columns dialysis removing unreacted agents present potential issues requiring attention include avoidance buffers containing primary amines(tris,glycine );maintaining environment devoid reducers necessary membrane protein optimization surfactant types/concentrations confirm whether targeted ionization efficiencies impacted prior mass spectral evaluations suggest running control experiments alongside gradient marked test sets determining optimal labelling parameters established best practices observed laboratory settings thus far have yielded promising results across multiple disciplines leveraging versatility exhibited throughout diverse scientific inquiries undertaken so far demonstrating tremendous promise ahead moving forward regarding future innovations surrounding related derivatives comparative technical advancements seen overtime resulting being prominent members belonging wider family comprising numerous variants existing today offering distinct characteristics suited differing requirements encountered daily basis laboratories worldwide driving further exploration into new avenues unexplored previously witnessed firsthand already paving way next generation breakthroughs unfolding rapidly all sectors alike pushing boundaries knowledge acquisition continuously evolving landscape surrounding contemporary sciences engaging minds brightening prospects awaiting just beyond horizon waiting unveil themselves fully fruition come life ultimately transforming how we perceive world itself entirely different perspectives offered unto us now never imagined possible beforehand altogether culminating exciting times lie ahead full realization possibilities abound infinitely out there yet remain untapped just waiting grasped fully understanding them thoroughly achieved collaboratively together!