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UltraBio™ Ni-NTA 琼脂糖磁珠 (His tag) 是一款金属螯合亲和磁胶，主要用于快速筛选和纯化带组氨酸标签 (His-tag) 的蛋白。其配基为次氮基三乙酸 (NTA)，配位金属离子为 Ni²⁺，琼脂糖磁珠是含顺磁性材料的琼脂糖凝胶微球。
与对应的柱层析琼脂糖填料 (Ni-NTA 亲和层析介质 6FF (His tag)，目录号为 N1524469) 相比，本产品拥有相同的配基修饰工艺，且保持了亲和力 (载量) 和特异性 (选择性) 之间良好的均衡：对大部分带组氨酸标签 (His tag) 蛋白的结合量大于 40 mg/mL 的同时，一步纯化后样品纯度可达 90%以上，在洗脱液中 Ni²⁺ 脱落率低，Ni²⁺与目标蛋白的摩尔比小于 0.1。
本产品的 NTA 配基与 Ni²⁺ 形成的四配位结构稳定性好，螯合后可耐受一定浓度的螯合剂、还原剂、碱性。样品中含有 1 mM EDTA、1~10 mM β-巯基乙醇或 5mM DTT时，可用 UltraBio™ Ni-NTA 琼脂糖磁珠 (His tag) 正常纯化，纯化后可用 0.1~1 M NaOH 清洗。
与琼脂糖填料的柱层析操作相比，除了保留 Ni-NTA 配基对带 Hig-tag 蛋白亲和层析高纯度、高收率、高重现性等特点外，利用磁胶的纯化操作还有以下特点和优势：
⁕ 料液一般无需澄清处理，可缩短纯化时间，对低表达量料液的富集效果更好；
⁕ 可进行手动或者自动化的批量操作，手动操作不依赖于设备，操作通量高。
本产品适用于带 His-tag 标签蛋白的小规模纯化与富集 (例如百毫克级)，高通量实验则可以快速筛选纯化条件和优化结果，且磁胶可以重复使用和用于放大生产过程。
包装规格： 20% v/v悬液
产品特点与技术指标：
2. 结合量：对大部分带组氨酸标签蛋白的结合量大于 40 mg/mL。 |
金属螯合磁胶亲和纯化原理简析：
金属离子亲和层析是基于蛋白质分子表面的组氨酸的咪唑基、半胱氨酸的巯基和色氨酸吲哚基能与 Cu²⁺、Ni²⁺、Zn²⁺、Co²⁺和 Ca²⁺形成稳定的配位结合而进行生物大分子分离纯化的过程。其中，以 Ni²⁺为螯合离子来纯化 6 个组氨酸组成的标签蛋白最为常见，本说明书也以该体系为例简要阐述 UltraBio™ Ni-NTA 琼脂糖磁珠 (His tag) 的亲和原理 (图 1)。
琼脂糖磁珠基球修饰上 NTA 配基后，该配体拥有四个配位基团，其中一个氮原子和三个氧原子可以与 Ni²⁺形成牢固的螯合物。Ni²⁺可形成 6 配位数的八面体结构，剩余的两个自由配位点可以与溶剂分子或蛋白质分子结合。Ni²⁺与组氨酸标签蛋白的两个组氨酸残基的咪唑基团配位结合的过程即为金属螯合配基与目标蛋白的特异性结合过程。提高缓冲液中咪唑的浓度可以将目标蛋白洗脱，洗脱过程中咪唑和目标蛋白竞争与 Ni²⁺的结合。降低 pH 会影响金属离子与配体的结合，因此改变 pH 也是金属螯合层析洗脱过程的方式之一，但 pH 不宜低于 4，pH 过低会将剥离金属离子。
金属离子亲和层析过程的非特异性吸附可能会来自离子相互作用、金属离子与含组氨酸、半胱氨酸、色氨酸蛋白之间的相互作用。前者通过提高缓冲液的离子强度得到抑制，通常添加 500 mM 的 NaCl；后者可在上样缓冲液中添加少量的咪唑 (例如~5 mM) 或者在冲洗过程中添加咪唑而除去。经过优化，可以得到样品纯度和收率均较高的结果。
UltraBio™ Ni-NTA 琼脂糖磁珠 (His tag) 磁胶在使用过程中 Ni²⁺掉落量少，可连续使用多次。当磁胶性能明显下降后可以用 EDTA 将原金属离子剥离，重新螯合镍后再生使用。
Ni-NTA金属螯合配基 与His标签蛋白特异性结合 咪唑竞争洗脱
图 1. UltraBio™ Ni-NTA 琼脂糖磁珠 (His tag) 金属螯合磁胶的纯化原理示意图
使用方法参考：
1. 磁胶准备与实验器材
① 计算磁胶用量
原包装是 20% 的磁胶分散液，可充分混匀后立即用移液枪按体积量取磁胶，每次量取均要重新混匀。50μL 分散液 (含10μL磁胶) 即可满足一次纯化。也可以通过沉降定量磁胶体积，沉降体积是指磁胶在纯水中自然沉降完全读取的稳定体积，沉降完全的时间通常需要 2h 以上，称取沉降后的磁胶 (上层不含保存液)，按 1g 磁胶≈1ml 磁胶称取，该方法更加适合纯化量较大的使用场景。通常将磁胶转移至 0.2~50 mL 规格的离心管中使用。
② 纯化所需的实验器材
手动操作时，利用磁胶纯化所需的主要器材涉及离心管和磁力架，市面上有适用于0.2~50 mL 离心管的不同形式的磁力架。磁性微球可在 5 秒内被吸至磁力架含磁铁的一侧，其余溶液则变澄清，形成固液的快速磁性分离，通过移液枪可方便移去清液。取出离心管，加入其他溶液后磁性微球易再分散。
利用磁胶纯化时，磁胶用量、料液体积与离心管规格的匹配程度比较关键。2mL 的离心管为例，磁胶的量控制在 10~100μL，样品料液的量控制在 0.3~1.5mL (离心管容积的 15%~75%)，料液与磁胶的体积比在 15~50 之间比较合适。使用其他规格的离心管时，可按比例变化。
③ 去除磁胶保存液
原磁胶保存液含 20%乙醇，使用前需要去除。量取磁胶后，加入离心管，在磁力架中使磁胶颗粒分离，用移液枪取出清夜。再向离心管中加入 10~50个磁胶体积的纯化水或者平衡缓冲液，盖紧盖子，手动震荡或者旋匀仪混合 10~30 秒使磁胶颗粒再分散，置于磁力架中，移去清液以除去保存液，重复操作 2~3 次。手动批量操作时可以将离心管置于离心管架，反复倒置达到再分散、混匀的目的。
2. 推荐缓冲液
金属螯合磁胶与磷酸盐、Tris-HCl 等缓冲体系兼容，其中 20 mM PB+150~500 mM NaCl (pH=7.4) 的体系最为常用。初始的缓冲液称为平衡缓冲液，记为缓冲液 A。对于 Ni-NTA 配基，缓冲液 A 可不含或者含 5~40 mM 咪唑，pH 在 7~8 之间居多，呈弱碱性。往 Buffer A 中添加 50~500 mM 的咪唑即为洗脱缓冲液，记为缓冲液 B，举例如下表。
| 平衡缓冲液 (缓冲液 A) | 20 mM PB+150 mM NaCl, pH=7.4 |
| 洗脱缓冲液 (缓冲液 B) | 20 mM PB+150 mM NaCl+500 mM 咪唑, pH=7.4 |
3. 平衡、孵育与淋洗
以 250μL 磁胶分散液 (含 50 μL 磁胶) 和 2mL 离心管的使用为例，介绍利用磁胶纯化带组氨酸的标签蛋白的过程。
① 平衡
按 1 小节的方法获取 50μL 磁胶于 2mL 离心管中，去除保存液。加入 500μL 缓冲液 A，在无磁场的条件下再分散磁胶 (手动震荡或者旋匀仪混合 10~30 秒)，将离心管置于在磁力架中，磁性分离后用移液枪移去平衡液。平衡过程可进行 1~2 次。
② 孵育
在上述移去平衡液后，往离心管里加入 1.5mL 样品料液，开始孵育。孵育过程中，磁胶颗粒需要保持均匀分散，可在旋匀仪或者台式摇床中进行。孵育通常在室温条件下进行，孵育时间在 30~60min 为宜，一些特殊样品也可以在 4 ℃的低温条件下孵育过夜。如果料液/磁胶体积比过大或者料液浓度过低，孵育时间可适当延长。料液可以不进行澄清处理，但需要保证在下述的淋洗过程中将颗粒物杂质除掉。通常情况下需要保留少量的料液以备 SDS-PAGE 等检测操作。孵育结束后将离心管置于磁力架上进行磁性分离，移出上清液到新的离心管中以备后续检测。
上样量可按 “mg 目标蛋白/mL 磁胶” 来设定，通常为磁胶结合量的 50~80%，例如 UltraBio™ Ni-NTA 琼脂糖磁珠 (His tag) 产品对重组链球菌蛋白 G的结合量为~40 mg/mL，纯化时的上样量可为 20~32 mg/mL。过载上样则可以用量评估磁胶的结合能力，例如 UltraBio™ Ni-NTA 琼脂糖磁珠 (His tag) 对不同组氨酸标签蛋白的结合能力。
③淋洗
往离心管加入 500μL 缓冲液 A，盖紧盖子，手动震荡或者旋匀仪混合 10~30 秒使磁胶颗粒再分散。将离心管置于磁力架上，磁性分离，待溶液变澄清后，移去上清液 (需取样检测可收集)。淋洗过程可进行 1~2 次，每次淋洗 操作耗时 2 min 左右。
4. 洗脱
最常用的洗脱方式为提高洗脱液中咪唑的浓度 (在缓冲液 A 中加入咪唑)，推荐在0~500mM 范围内进行阶跃洗脱或者线性梯度洗脱，确定最优的洗脱浓度。对于大多数带组氨酸标签的蛋白，500 mM 的咪唑浓度可以将目标蛋白洗脱完全。
往离心管中加入 5~10 体积的缓冲液 B，即 250~500μL，盖紧盖子，手动震荡或者旋匀仪混合 10~30 秒使磁胶颗粒再分散。将离心管置于磁力架上，磁性分离，待溶液变澄清后，收集洗脱液，洗脱过程可进行 1~3 次。如果认为洗脱过程需要孵育，孵育时间控制在 10min 为宜。
5. 再生与在位清洗
如果使用目的为小批量纯化，例如用 50μL 磁胶纯化 1mg 目标蛋白的过程，磁胶在再生与清洗后可重复使用。如果使用目的为高通量的筛选和优化，磁胶通常是一次性使用。
① 再生
从磁力架上取下离心管，加入 500μL 缓冲液 A，盖紧盖子，手动震荡或者旋匀仪混合 10~30 秒使磁胶颗粒再分散，将离心管置于磁力架上，进行磁性分离，待溶液变澄清后，移去上清液，再生过程可进行 1~2 次。
② CIP
通常上述的再生过程可将磁胶彻底清洗。若有蛋白与磁胶强烈结合无法洗脱，或有沉淀、变性蛋白时，需要对磁胶进行在位清洗，可采用以下方法去除杂质。
| CIP 过程 | 目的 |
| 2 M NaCl，3~5 CV；水，5~10 CV×2。 | 除去通过离子相互作用结合的杂质 |
| 0.5 M NaOH，3~5 CV，1~2 h 的接触时间，若除去内毒素则需更长时间；缓冲液 A，5~10 CV；水，5~10 CV×2。 | 去除蛋白沉淀、疏水性蛋白或去除热源等 其他杂质 |
| 30%异丙醇或 70%乙醇，5~10 CV；水，5~10 CV×2。 | 去除紧密结合的脂蛋白和脂质体等 |
6. 筛选与优化
上述纯化条件适用于大部分带组氨酸标签蛋白的纯化过程，当需要得到最佳的使用条件，或者为对应的填料筛选与优化纯化条件时，可从以下几个方面考虑：缓冲体系，特别是洗杂/洗脱过程的咪唑浓度；孵育的条件，例如温度与时间；料液/磁胶的体积比。
7. 磁胶保存
磁胶的初始保存液为 20%乙醇，使用过后可继续用 20%乙醇保存。保存温度在 2~30 ℃为宜，不可冻存。
UltraBio™ Ni-NTA Magnetic Agarose Beads (His tag) are metal‑chelating affinity magnetic beads designed for rapid screening and purification of histidine‑tagged (His‑tag) proteins. The ligand is nitrilotriacetic acid (NTA), with Ni²⁺ as the coordinated metal ion. The agarose magnetic beads are agarose‑gel microspheres incorporated with paramagnetic material.
Compared with the corresponding column‑chromatography agarose resin (Ni-NTA Sepharose 6FF (His tag), Catalog No. N1524469), this product adopts an identical ligand‑modification process and maintains an excellent balance between binding capacity (loading capacity) and specificity (selectivity). It exhibits a binding capacity of more than 40 mg/mL for most His‑tagged proteins, and sample purity above 90 % can be achieved after one‑step purification. The leakage rate of Ni²⁺ in the eluate is low, and the molar ratio of stripped Ni²⁺ to target protein is less than 0.1.
The tetra‑coordination structure formed between the NTA ligand and immobilized Ni²⁺ is highly stable. After chelation, the resin tolerates certain concentrations of chelating agents, reducing agents and alkaline conditions. UltraBio™ Ni-NTA Magnetic Agarose Beads (His tag) can be used for normal purification when samples contain 1 mM EDTA, 1‑10 mM β‑mercaptoethanol or 5 mM DTT. The beads can be cleaned with 0.1‑1 M NaOH after purification.
In comparison with column‑chromatography workflows using agarose resin, this magnetic‑bead‑based purification retains the advantages of Ni‑NTA ligands for His‑tag protein affinity chromatography, including high purity, high yield and excellent reproducibility. In addition, the magnetic‑bead‑based purification workflow features the following benefits:
⁕ Feedstock solutions generally do not require clarification, which shortens purification time and delivers superior enrichment performance for samples with low‑level protein expression;
⁕ Manual or automated batch processing is supported. Manual operations require no specialized equipment and enable high‑throughput sample‑processing capacity.
This product is suitable for small‑scale purification and enrichment (e.g., hundreds‑of‑milligram scale) of His‑tagged proteins. It enables fast screening and optimization of purification conditions for high‑throughput experiments. Furthermore, the magnetic beads are reusable and can be applied to scale‑up production processes.
Packaging specification: 20% (v/v) suspension.
Product Features and Technical Specifications:
2. Binding capacity: More than 40 mg/mL for most histidine‑tagged proteins. |
Principle of Affinity Purification Using Metal‑Chelating Magnetic Beads:
Immobilized‑metal‑affinity chromatography (IMAC) is a separation‑purification technique for biomacromolecules. It relies on stable coordinate bonds formed between the imidazole group of surface‑exposed histidine residues, thiol group of cysteine residues, indole group of tryptophan residues on proteins and metal ions including Cu²⁺, Ni²⁺, Zn²⁺, Co²⁺ and Ca²⁺. Purification of 6‑histidine‑tagged proteins using Ni²⁺ as the chelated metal ion is the most‑common application. Taking this system as an example, this manual briefly describes the affinity principle of UltraBio™ Ni-NTA Magnetic Agarose Beads (His tag) (Figure 1).
After the base Magnetic Agarose Beads are modified with NTA ligands, each ligand contains four coordinating groups. One nitrogen atom and three oxygen‑atom sites form a stable chelate complex with Ni²⁺. Ni²⁺ adopts an octahedral geometry with a coordination number of six; the remaining two free coordination sites are accessible to solvent molecules or target‑protein molecules. The specific binding between the metal‑chelating ligand and the target protein occurs when Ni²⁺ coordinates with the imidazole groups of two histidine residues on the His‑tagged protein. The target protein can be eluted by increasing the imidazole concentration in the buffer, since imidazole competes with the target protein for binding sites on Ni²⁺. A reduction in pH weakens the interaction between the metal ion and the ligand. Therefore, pH adjustment serves as an alternative elution strategy for immobilized‑metal‑affinity‑chromatography. However, the pH should not drop below 4, because excessively low pH will strip the metal ion from the resin.
Non‑specific adsorption during immobilized‑metal‑affinity‑chromatography (IMAC) may arise from ionic interactions, as well as interactions between metal ions and endogenous proteins containing histidine, cysteine or tryptophan residues. The former can be suppressed by increasing the ionic strength of the buffer, typically by adding 500 mM NaCl. The latter can be eliminated by adding a small amount of imidazole (e.g. ~5 mM) to the loading buffer or including imidazole in the wash buffer. After optimization, both high sample purity and high yield can be achieved.
UltraBio™ Ni-NTA Magnetic Agarose Beads (His tag) exhibit low Ni²⁺ leakage during operation and can be reused for multiple cycles. When the performance of the magnetic beads decreases significantly, the original metal ions can be stripped off with EDTA. The resin can then be regenerated by re‑chelating nickel ions for subsequent use.
Ni‑NTA metal‑chelating ligand Specific binding to His‑tagged protein Imidazole competitive elution
Figure 1. Schematic diagram of purification principle for UltraBio™ Ni-NTA Magnetic Agarose Beads (His tag) metal‑chelating magnetic beads
Reference Protocol:
1. Magnetic‑bead Preparation and Experimental Supplies
① Calculation of magnetic‑bead dosage
The original product is supplied as a 20 % magnetic‑bead dispersion. Mix thoroughly immediately before pipetting the required volume. Re‑homogenize the suspension before every sampling.
50 μL of dispersion (containing 10 μL settled magnetic beads) is sufficient for one‑run purification. Magnetic‑bead volume can also be quantified by sedimentation. The settled volume refers to the stable volume read‑out after complete sedimentation of beads in pure water, which usually takes no less than 2 hours. Weigh the settled beads (without supernatant storage solution). 1 g of settled beads ≈ 1 mL bead volume. This gravimetric method is more suitable for large‑scale purification. Generally, transfer the beads into centrifuge tubes ranging from 0.2 mL to 50 mL for experiments.
② Experimental equipment for purification
For manual purification, centrifuge tubes and a magnetic rack are the main required supplies. Commercial magnetic racks compatible with 0.2‑50 mL centrifuge tubes are available. Magnetic microspheres can be attracted to the magnet‑containing side of the rack within 5 seconds, leaving a clear supernatant and achieving rapid solid‑liquid magnetic separation. The supernatant can then be conveniently removed by pipette. After taking the tube off the rack and adding new solution, the magnetic beads can be easily resuspended. Matching the bead dosage, feed‑solution volume and tube size is critical.
Take a 2 mL centrifuge tube as an example: use 10‑100 μL settled beads and 0.3‑1.5 mL feed solution (15 %‑75 % of tube volume). A volume ratio between feed solution and settled beads of 15‑50 is recommended. Scale the volumes proportionally for other tube sizes.
③ Removal of bead‑storage solution
The original storage solution contains 20 % ethanol and must be removed before use. After pipetting beads into the centrifuge tube, place the tube on the magnetic rack to pellet the beads and aspirate off the supernatant. Add purified water or binding buffer at 10‑50‑fold the settled‑bead volume, cap tightly, and resuspend the beads by manual vortexing or mixer for 10‑30 seconds. Place the tube back on the magnetic rack and discard the supernatant to remove residual storage solution. Repeat this washing step 2‑3 times. For manual batch processing, resuspension and mixing can be achieved by repeated inversion of tubes placed in a tube rack.
2. Recommended Buffers
Metal‑chelating magnetic beads are compatible with buffer systems such as phosphate and Tris‑HCl. Among them, 20 mM PB + 150‑500 mM NaCl (pH = 7.4) is the most commonly‑used system. The initial buffer is referred to as the binding buffer, designated Buffer A. For Ni‑NTA ligands, Buffer A may contain no imidazole or 5‑40 mM imidazole, with a weakly‑alkaline pH typically ranging from 7 to 8. The elution buffer, designated Buffer B, is prepared by adding 50‑500 mM imidazole to Buffer A. Typical examples are shown in the table below:
| Binding buffer (Buffer A) | 20 mM PB+150 mM NaCl, pH=7.4 |
| Elution buffer (Buffer B) | 20 mM PB + 150 mM NaCl + 500 mM imidazole, pH = 7.4 |
3. Equilibration, Incubation and Washing
The purification workflow for histidine‑tagged protein is described below using 250 μL magnetic‑bead dispersion (containing 50 μL settled beads) and a 2 mL centrifuge tube as an example.
① Equilibration
Transfer 50 μL settled magnetic beads into a 2 mL centrifuge tube and remove the storage solution following the procedure described in Section 1. Add 500 μL Buffer A and resuspend the beads thoroughly in the absence of a magnetic field (vortex manually or mix on a rotator for 10‑30 seconds). Place the tube on a magnetic rack. After magnetic separation, aspirate and discard the equilibration buffer. The equilibration step may be repeated 1‑2 times.
② Incubation
After removal of the equilibration buffer, add 1.5 mL sample feed solution to the tube to start incubation. The magnetic beads must remain evenly dispersed throughout incubation. Mixing can be performed on a rotator or benchtop shaker. Incubation is generally carried out at room temperature for 30‑60 min. Certain sensitive samples may be incubated overnight at 4 °C. Incubation time can be extended appropriately if the volume ratio of feed solution to beads is too high or the protein concentration is very low. The feed solution does not require pre‑clarification; however, particulate contaminants must be removed in the subsequent washing step. Retain an aliquot of the feed solution for downstream analysis such as SDS‑PAGE. Upon completion of incubation, place the tube on the magnetic rack for magnetic separation. Transfer the supernatant to a new centrifuge tube for further detection.
Sample loading is calculated in mg target protein per mL settled beads. A loading of 50‑80 % of the bead binding capacity is recommended. For instance, UltraBio™ Ni-NTA Magnetic Agarose Beads (His tag) exhibit a binding capacity of ~40 mg/mL for recombinant Protein G from Streptococcus, and a loading amount of 20‑32 mg/mL can be applied during purification. Over‑loading can be used to evaluate the actual binding capacity of the magnetic beads toward different His‑tagged proteins.
③ Washing
Add 500 μL Buffer A to the centrifuge tube, cap tightly, and resuspend the beads by manual shaking or mixing on a rotator for 10‑30 seconds. Place the tube on the magnetic rack for magnetic separation. Once the supernatant becomes clear, aspirate and discard it (collect the supernatant if sampling‑for‑assay is required). Repeat the washing step 1‑2 times, with approximately 2 min incubation for each wash.
4. Elution
The most‑frequently‑used elution strategy is to increase the imidazole concentration in the eluent (imidazole supplemented into Buffer A). Step‑wise elution or linear‑gradient elution within the range of 0‑500 mM is recommended to determine the optimal elution concentration. For most histidine‑tagged proteins, 500 mM imidazole is sufficient for complete elution of the target protein.
Add 5‑10 bead volumes of Buffer B, i.e. 250‑500 μL, to the centrifuge tube. Cap tightly and resuspend the magnetic beads by manual shaking or mixing on a rotator for 10‑30 seconds. Place the tube on a magnetic rack for magnetic separation. Collect the eluate once the supernatant turns clear. The elution step may be repeated 1‑3 times. If incubation during elution is required, an incubation time of 10 min is recommended.
5. Regeneration and Clean‑in‑Place (CIP)
For small‑scale purification workflows, e.g. purification of 1 mg target protein using 50 μL settled magnetic beads, the beads can be reused after regeneration and cleaning. For high‑throughput screening and optimization experiments, the magnetic beads are generally intended for single‑use only.
① Regeneration
Remove the centrifuge tube from the magnetic rack, add 500 μL Buffer A and cap tightly. Resuspend the magnetic beads by manual shaking or mixing on a rotator for 10‑30 seconds. Place the tube back on the magnetic rack for magnetic separation. Discard the supernatant once the solution becomes clear. The regeneration step may be repeated 1‑2 times.
② Clean‑in‑Place (CIP)
The regeneration procedure described above is usually sufficient for thorough cleaning of the beads. If tightly‑bound protein cannot be eluted, or precipitates / denatured proteins remain on the beads, clean‑in‑place treatment shall be performed to remove contaminants using the protocols
below:
| CIP Procedure | Purpose |
| 2 M NaCl, 3‑5 CVPurified water, 5‑10 CV, twice | Removal of contaminants bound via ionic interactions |
| 0.5 M NaOH, 3‑5 CV; contact time 1‑2 h (extended contact time is required for endotoxin removal)Buffer A, 5‑10 CVPurified water, 5‑10 CV, twice | Removal of protein precipitates, hydrophobic proteins, endotoxins (pyrogens) and other impurities |
| 30 % isopropanol or 70 % ethanol, 5‑10 CVPurified water, 5‑10 CV, twice | Removal of tightly‑bound lipoproteins, liposomes and similar substances |
6. Screening and Optimization
The purification‑conditions described above are suitable for most histidine‑tagged proteins. To achieve optimal performance, or to screen and optimize purification parameters for the resin, the following factors can be evaluated: buffer composition, especially imidazole concentrations used in the wash‑step and elution‑step; incubation parameters such as temperature and duration; volume ratio of feed‑solution to settled magnetic beads.
7. Magnetic‑Bead Storage
The original storage solution for the magnetic beads is 20 % ethanol. After use, the beads can be stored again in 20 % ethanol. The recommended storage temperature range is 2‑30 °C. Do not freeze.
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