储存缓冲液：20mM Tris, 0.01% Tween-20, 0.05% KroVin300, pH7.4
UltraBio™ 链霉亲和素磁珠 (1μm) 是将链霉亲和素（Streptavidin）通过化学方法固定在聚合物磁性微球（MagPoly Beads）上，应用生物素与链霉亲和素配体之间的相互作用来实现固定化生物素或生物素化的蛋白、抗体等物质的目的。
参数
| 指标
|
基质
| 聚合物磁性微球 |
配基
| 链霉亲和素 |
| 磁珠浓度 | 10 mg/mL
|
粒径
| 1 μm |
载量
| ＞20 μg Biotinylated IgG/mg磁珠
|
保存
| 2-8℃ (20mM Tris, 0.01% Tween-20, 0.05% KroVin300, pH7.4） |
使用说明：
本操作流程以每次反应使用100μl UltraBio™ 链霉亲和素磁珠 (1μm) 为例，可根据需要适当的增加或减少磁珠使用量。
一、试剂准备
1、核酸样本（Buffer Ⅰ）：10mM Tris-HCl (pH7.5), 1mM EDTA, 1M NaCl, 0.01%~0.1% Tween-20；
2、抗体/蛋白样本（Buffer Ⅱ）：1×PBS, pH7.4, 含0.05% Tween-20，可根据需要添加0.01%~0.1% BSA。
注：以上为推荐使用的缓冲液成分，用户可根据具体实验需求调整缓冲液盐浓度及pH。
二、生物素化分子的捕获
1、核酸的捕获
(1) 将链霉亲和素磁性微球颠倒或漩涡振荡，混合均匀。
(2) 取 100μl 链霉亲和素磁性微球加入新的离心管中。放置在磁分离器上，待溶液变澄清后，用移液器吸弃保护液。
(3) 取下离心管加入1ml BufferⅠ，充分振荡重悬磁珠，混匀后放置在磁分离器上，待溶液变澄清后，用移液器吸弃上清，收集磁珠，重复洗3次。
注：当步骤(2)中取用磁珠体积大于1ml时，加入与磁珠体积相同的BufferⅠ。
(4) 加入 500μl 处理好的核酸样本（核酸样本为 BufferⅠ 稀释好的生物素化核酸）使磁珠浓度为 2mg/ml，充分振荡重悬磁珠，可将离心管置于旋转混合仪上，室温旋转混合30min，混合时间可根据实际情况进行调整。
(5) 混匀后将离心管放置在磁分离器上，待溶液变澄清后，用移液器吸弃上清，建议将上清液转移至新的离心管，用于后续计算结合到磁珠上的核酸量。按步骤 (3)的方法洗涤磁珠，重复洗3次。
(6) 根据后续实验要求，加入合适的低盐缓冲液，重悬磁珠。至此，结合生物素化核酸步骤完成，磁珠可用于后续操作。
(7) 结合到磁珠上的核酸量可根据测定反应前后核酸的浓度计算：核酸量=（反应前浓度-反应后浓度）× 反应溶液。
2、抗体/蛋白的捕获
(1) 将链霉亲和素磁性微球颠倒或漩涡振荡，混合均匀。
(2) 取 100μl 链霉亲和素磁性微球加入新的离心管中。放置在磁分离器上，待溶液变澄清后，用移液器吸弃保护液。
(3) 取下离心管加入1ml Buffer Ⅱ，充分振荡重悬磁珠，混匀后放置在磁分离器上，待溶液变澄清后，用移液器吸弃上清，收集磁珠，重复洗 3 次。
注：当步骤(2)中取用磁珠体积大于1ml 时，加入与磁珠体积相同的Buffer Ⅱ。
(4) 加入1ml 处理好的抗体/蛋白样本（抗体/蛋白样本为 Buffer Ⅱ稀释好的生物素化抗体/蛋白）使磁珠浓度为1mg/ml，充分振荡重悬磁珠，可将离心管置于旋转混合仪上，室温旋转混合 60min，混合时间可根据实际情况进行调整。
(5) 混匀后将离心管放置在磁分离器上，待溶液变澄清后，用移液器吸弃上清，建议将上清液转移至新的离心管，用于后续计算结合到磁珠上的抗体/蛋白量。按步骤(3)的方法洗涤磁珠，重复洗 5 次。
(6) 根据后续实验要求，加入 Buffer Ⅱ或其他合适的缓冲液，重悬磁珠。至此，结合生物素化抗体/蛋白步骤完成，磁珠可用于后续操作。
(7) 结合到磁珠上的抗体/蛋白量可根据测定反应前后抗体/蛋白的浓度计算：抗体/蛋白量=（反应前浓度-反应后浓度）×反应溶液体积。
注意事项：
1、避免对磁珠进行冷冻等操作。
2、从磁珠保存管中移取磁珠前应充分振荡重悬均匀，操作过程中应避免产生气泡。
3、因不同项目存在差异化，应根据实验需求，优化具体使用参数（包括时间、缓冲液、载量等）。
UltraBio™ Streptavidin Magnetic Beads (1 μm) are prepared by chemically immobilizing streptavidin onto polymeric magnetic microspheres (MagPoly Beads). Utilizing the interaction between biotin and streptavidin ligands, these beads can capture immobilized biotin or biotinylated substances such as proteins and antibodies.
Parameter
| Indicator
|
Matrix
| Polymeric magnetic microspheres
|
Ligand
| Streptavidin
|
Bead Concentration
| 10 mg/mL
|
Particle Size
| 1 μm |
Binding Capacity
| ＞20 μg biotinylated IgG per mg beads
|
Storage
| 2-8℃ (20mM Tris, 0.01% Tween-20, 0.05% KroVin300, pH7.4） |
Instructions for Use:
The following protocol takes 100 μL of UltraBio™ Streptavidin Magnetic Beads (1 μm) per reaction as an example. The bead dosage can be increased or decreased as required.
ⅠReagent Preparation
1. Nucleic acid sample (Buffer Ⅰ): 10 mM Tris-HCl (pH 7.5), 1 mM EDTA, 1 M NaCl, 0.01%~0.1% Tween-20
2. Antibody/protein sample (Buffer Ⅱ): 1× PBS (pH 7.4) containing 0.05% Tween-20. 0.01%~0.1% BSA may be added if needed.
Note: The above are recommended buffer formulations. Adjust the salt concentration and pH according to specific experimental requirements.
ⅡCapture of Biotinylated Molecules
1. Capture of Nucleic Acids
① Invert or vortex the streptavidin magnetic beads to mix thoroughly.
② Transfer 100 μL beads into a new centrifuge tube. Place the tube on a magnetic separator. Once the solution turns clear, aspirate and discard the storage solution with a pipette.
③ Remove the tube from the separator, add 1 mL Buffer Ⅰ, and vortex fully to resuspend the beads. Place the tube back on the magnetic separator until the solution is clear, then aspirate the supernatant. Repeat the washing step 3 times.
Note: If the bead volume taken in Step (2) exceeds 1 mL, add Buffer Ⅰ of the same volume as the beads.
④ Add 500 μL prepared nucleic acid sample (biotinylated nucleic acid diluted with Buffer Ⅰ) to reach a bead concentration of 2 mg/mL. Vortex to fully resuspend the beads. Place the tube on a rotator and incubate with rotation at room temperature for 30 minutes. The incubation time can be adjusted as needed.
⑤ After incubation, place the tube on the magnetic separator and wait for the solution to clear. Aspirate the supernatant, which is recommended to be transferred to a new tube for subsequent quantification of bound nucleic acids. Wash the beads 3 times following the procedure in Step (3).
⑥ Resuspend the beads with an appropriate low-salt buffer based on downstream experimental requirements. The capture of biotinylated nucleic acids is now complete, and the beads are ready for subsequent procedures.
⑦ The amount of bound nucleic acids is calculated as follows:
Bound nucleic acid mass = (Initial concentration − Final concentration) × Total reaction volume
2. Capture of Antibodies/Proteins
① Invert or vortex the streptavidin magnetic beads to mix thoroughly.
② Transfer 100 μL beads into a new centrifuge tube. Place the tube on a magnetic separator. Once the solution turns clear, aspirate and discard the storage solution with a pipette.
③ Remove the tube from the separator, add 1 mL Buffer Ⅱ, and vortex fully to resuspend the beads. Place the tube back on the magnetic separator until the solution is clear, then aspirate the supernatant. Repeat the washing step 3 times.
Note: If the bead volume taken in Step (2) exceeds 1 mL, add Buffer Ⅱ of the same volume as the beads.
④ Add 1 mL prepared antibody/protein sample (biotinylated antibody/protein diluted with Buffer Ⅱ) to reach a bead concentration of 1 mg/mL. Vortex to fully resuspend the beads. Place the tube on a rotator and incubate with rotation at room temperature for 60 minutes. The incubation time can be adjusted as needed.
⑤ After incubation, place the tube on the magnetic separator and wait for the solution to clear. Aspirate the supernatant, which is recommended to be transferred to a new tube for subsequent quantification of bound antibodies/proteins. Wash the beads 5 times following the procedure in Step (3).
⑥ Resuspend the beads with Buffer Ⅱ or other suitable buffers based on downstream experimental requirements. The capture of biotinylated antibodies/proteins is now complete, and the beads are ready for subsequent procedures.
⑦ The amount of bound antibodies/proteins is calculated as follows:
Bound antibody/protein mass = (Initial concentration − Final concentration) × Total reaction volume
Precautions:
1. Do not freeze the magnetic beads.
2. Fully resuspend and homogenize the beads before pipetting. Avoid bubble generation during operation.
3. Optimize experimental parameters (including incubation time, buffer formulation, binding capacity, etc.) according to different experimental applications.
ZJ26F1039864
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