Percoll PLUS/Percoll
CELL PREPARATION

Percoll™ PLUS is a silica-based colloidal medium for cell separation by density gradient centrifugation. The silica particles of the medium are covalently coated with silane, providing product stability and long shelf life. The silane coating also provides low osmolality and toxicity, as well as low viscosity. Percoll PLUS has low levels of endotoxins, making it well-suited for cell separation
in clinical research applications. Cell separation with the medium is performed under gentle conditions, facilitating the isolation
of a variety of cells, subcellular particles, and viruses where preservation of viability and morphological integrity is important. The low toxicity of Percoll PLUS ensures that removal of the medium from separated cellular particles is not usually necessary.
After adjustment, Percoll PLUS forms iso-osmotic gradients within the density range of 1.0 to 1.3 g/ml. This density range is especially useful since most cells, subcellular particles, and
viruses have a buoyant density of 1.0 to 1.2 g/ml in Percoll PLUS.

Percoll PLUS offers:

1. Low endotoxin levels (max. 2 EU/ml)
2. Absence of toxicity for cells and very low chemical reactivity
3. Low osmolality: Percoll PLUS can easily be adjusted with physiological saline, other balanced salt solutions, or cell culture media, to give gradients that are iso-osmotic throughout
4. Low viscosity resulting in rapid formation of gradients and particle separation at low centrifugal forces

Specifications for Percoll PLUS are shown in Table 1.

Percoll PLUS gradients
Percoll PLUS can be used in density centrifugation applications for the isolation and purification of cells, subcellular particles, and viruses down to ~70S. The medium offers both high resolution and biological compatibility. The low intrinsic osmolality and viscosity are particularly useful when isolating organelles where maintenance of membrane integrity is important.

Fig 1. Percoll PLUS provides reliable separation of cells, subcellular particles, and viruses by density gradient centrifugation.
Table 1. Percoll PLUS specifications

Percoll PLUS can be used to produce preformed, continuous or discontinuous gradients. Under moderate centrifugal force, the colloidal particles that comprise Percoll PLUS sediment to form smooth, continuous density gradients. This property can be exploited in either fixed-angle or vertical rotors.
The medium is also suited to applications where high-speed centrifugation is required. In this case, the sample can be premixed with the medium and subsequently separated on the continuous gradient formed in situ. Thus, gradient formation and sample separation can be achieved in one step. All experiments described in the literature using Percoll can also be performed with Percoll PLUS.
 

Percoll

Percoll is a silica-based colloidal medium for cell separation by density gradient centrifugation (see Percoll specifications in Table 2). The silica particles of the medium are coated with polyvinylpyrrolidone (PVP), which gives low osmolality and low viscosity. Cell separation with Percoll is performed under gentle conditions, facilitating the isolation of a variety of cells,
subcellular particles, and viruses. Iso-osmotic Percoll gradients can be formed within the density range of 1.0 to 1.3 g/ml.
This density range is optimized for separation of most cells, subcellular particles, and larger viruses, which have a buoyant density of 1.0 to 1.2 g/ml in Percoll.

Percoll offers:

1. Low osmolality: Percoll can easily be adjusted with physiological saline, cell culture medium, or sucrose to give gradients that are iso-osmotic throughout
2. Low viscosity resulting in rapid formation of gradients and particle separation at low centrifugal forces
3. Support through extensive research use: Thousands of publications on Percoll in scientific journals
4. Formation of either continuous preformed or selfgenerated gradients by centrifugation at moderate speed

Table 2. Percoll specifications

密度：1.13g/ml

适合分离细胞，亚细胞成分和大病毒(>70S)，渗透压均匀，粘度低，可调至生理离子强度和pH，分离条件温和，对细胞无毒性，可以灭菌消毒。

Percoll分离液是一种经过聚乙烯吡咯烷酮处理过的硅胶颗粒，经过高速离心后可形成连续密度梯度，由于Percoll扩散常数低，所形成的梯度十分稳定。Percoll分离液采用预先形成的密度梯度时可在低离心力(200～1000g)于数分至数十分钟内达到满意的细胞分离结果。此外，Percoll分离液不穿透生物膜，对细胞无毒害，因此广泛用于分离细胞、亚细胞成分、细菌及病毒，还可将受损细胞及其碎片与完好的活细胞分离。Percoll常用于分离和传话植物原生质体、核、叶绿体、和线粒体。

Percoll分离液的配置与使用

1、不同浓度(密度)Percoll分离液的制备: 先用9份Percoll分离液与1份8.5％ NaCl混合达到生理性渗透压，然后用生理溶液(0.85％ NaCl)稀释到所需浓度。

2、不连续密度梯度Percoll层的制备: 先将试管壁用牛血清湿润，除去多余血清，这种预处理可使逐层叠加的Percoll液平稳沿管壁流下，使形成满意的界面。在制备过程中一般用长针头注射器从高密度向低密度逐层放置，有时相邻两层Percoll比重相差不大时，可将Percoll液放入注射器中，小针头斜面紧贴管壁，任其自然慢慢流下。

3、装样：样品体积和细胞浓度根据不同细胞而异，一般加样体积不宜过大，细胞浓度也不可过高，否则会影响细胞的分离和回收。

4、离心：一般采用离心力为400g，时间20～25min。由于多层Percoll之间密度差别不大，因此离心机加速、降速时要慢，要平稳。

5、取样：当所要分离的细胞绝大部分在两层的界面时，可逐层去除Percoll液后收集界面部位的细胞;有时大部分细胞位于Percoll层中,则需要逐层收集。收获含有Percoll液的细胞经2次洗涤后可供培养或检测用。

Percoll细胞分离液的优点

Percoll渗透性低、不穿透细胞、密度高、无毒害，是目前较理想的介质。Percoll密度梯度离心已被多次成功地用于动物细胞及其细胞器的分离，纯化了包括人血液细胞、骨髓细胞、红血球细胞、白血球细胞、淋巴细胞、肝细胞等在内的十余种动物细胞。

Percoll细胞分离液注意事项：

1、Percoll细胞分离液在储存及孵育过程中避免将实际保留在强光中。

2、试剂瓶盖需盖紧，防止蒸发和污染。