刺桐凝集素（ECL）结合的糖结构广泛存在于哺乳动物来源的膜糖蛋白与血清糖蛋白上。若该糖结构发生唾液酸化修饰，则会阻碍凝集素结合。利用这一特异性，可将固定于琼脂糖微球的 ECL 用于哺乳动物糖蛋白的分离与分级。已有文献报道，该凝集素可通过阴性选择铺板法分离人自然杀伤细胞（NK 细胞）。人 NK 细胞表面不存在可供 ECL 结合的糖结构；与其他单核细胞不同，NK 细胞不会粘附在包被 ECL 的培养皿上。该方法属于阴性选择铺板法，能够高效获得高存活率的 NK 细胞。被粘附捕获的细胞可通过半乳糖或乳糖孵育实现洗脱回收。生物素化刺桐凝集素的生物素标记数量适中，可赋予该凝集素最优染色性能。本偶联物基本不含游离生物素，叠氮化钠作为防腐剂。
技术信息
刺桐凝集素由两种亚基组成，分子量约 28 kDa 与 26 kDa。ECL 识别结合的糖结构普遍存在于哺乳动物膜糖蛋白及血清糖蛋白；该位点发生唾液酸化后将抑制凝集素结合。依托该特异性，琼脂糖固定化 ECL 可用于哺乳动物糖蛋白的分离、分级。
这款生物素化凝集素非常适合依托生物素 - 亲和素 / 链霉亲和素系统开展糖复合物检测。实验流程：先加入生物素化凝集素，随后配合亲和素 - 生物素复合物试剂、亲和素 D 偶联物或链霉亲和素衍生物进行检测。
抑制糖 / 洗脱糖：200 mM 乳糖
规格参数
分子量
| 54 kDa | 消光系数
| 1.3 | 缓冲配方
| 10 mM HEPES, 0.15 M NaCl, pH 7.5, 0.1 mM CaCl₂, 0.08% sodium azide. | 抑制 / 洗脱糖
| 乳糖 | 糖结合特异性
| 末端 2 型 LacNAc、末端 2 型 LacdiNAc
| 使用简述
| 多数实验体系，建议使用上述缓冲液新鲜配制浓度 5–20 µg/mL 的工作液。
| 应用
| Immunohistochemistry / Immunocytochemistry, Immunofluorescence, Blotting Applications, Elispot, ELISAs, Glycobiology
| 浓度
| 5 mg 活性偶联物 /mL
| 标记类型
| Biotinylated |
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产品常见问题
问：我采购了一款生物素化凝集素。产品说明书建议配制工作液时，缓冲液中添加 0.1 mM Ca²⁺。请问应当添加什么试剂，为何需要钙离子？
答：根据经验，部分凝集素需要钙离子（Ca²⁺）才能达到最佳结合活性。建议向工作液中补充氯化钙（CaCl₂），保证体系钙离子维持最低有效浓度。若选用磷酸盐缓冲液作为稀释液或储存液，该注意事项尤为重要。
The carbohydrate structure to which ECL binds is frequently found in membrane and serum glycoproteins of mammalian origin. Sialic acid substitution on this structure appears to prevent the lectin from binding. This specificity offers an opportunity to utilize agarose bound ECL to isolate or fractionate mammalian glycoproteins.
This lectin has been reported to be useful for the isolation of human natural killer (NK) cells using a negative selection panning technique. Human NK cells appear to lack accessible surface carbohydrate structures required for binding ECL and, unlike other mononuclear cells, do not adhere to ECL-coated culture dishes. Since this procedure involves a negative selection panning technique, a high recovery of viable NK cells can be obtained. The adherent cells can also be recovered by incubation in galactose or lactose.
Biotinylated Erythrina cristagalli lectin has an appropriate number of biotins bound to provide the optimum staining characteristics for this lectin. This conjugate is supplied essentially free of unconjugated biotins and is preserved with sodium azide.
Technical Information
Erythrina cristagalli lectin consists of two different subunits of approximately 28 kDa and 26 kDa. The carbohydrate structure to which ECL binds is frequently found in membrane and serum glycoproteins of mammalian origin. Sialic acid substitution on this structure appears to prevent the lectin from binding. This specificity offers an opportunity to utilize agarose bound ECL to isolate or fractionate mammalian glycoproteins.
This biotinylated lectin is an ideal intermediate for examining glycoconjugates using the Biotin-Avidin/Streptavidin System. First the biotinylated lectin is added, followed by the Avidin-Biotin Complex Reagent, Avidin D conjugate, or streptavidin derivative.
Inhibiting/Eluting Sugar: 200 mM lactose
Specifications
| Molecular Weight | 54 kDa
| | Extinction Coefficient | 1.3 | | Formulation | 10 mM HEPES, 0.15 M NaCl, pH 7.5, 0.1 mM CaCl₂, 0.08% sodium azide. | | Inhibiting or Eluting Sugar | Lactose | | Sugar Specificity | Terminal type 2 LacNAc, and terminal type 2 LacdiNAc | | Usage Summary | For most applications we recommend a freshly prepared working solution of 5–20 µg/ml in the below buffer. | | Applications | Immunohistochemistry / Immunocytochemistry, Immunofluorescence, Blotting Applications, Elispot, ELISAs, Glycobiology | | Concentration | 5 mg active conjugate/ml | | Conjugate | Biotinylated |
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Product FAQs
I recently purchased a biotinylated lectin. The datasheet supplied with the lectin suggests including 0.1 mM Ca²⁺ as part of the recommended buffer to prepare a working solution. What should I specifically add, and why is this required?
From our experience we have found that some lectins require Ca²⁺ to be present for optimal binding activity. We suggest using calcium chloride (CaCl₂) to fortify working solutions and ensure a minimum level of Ca²⁺ is met. This may be particularly pertinent if using phosphate-based buffers as diluents and storage solutions.
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