Specific Cell Characteristics

Cellular materials range in size from 1 |im to 50 |im. Cell populations are classified by a set of morphological, functional and biophysical characteristics. The biophysical characteristics are of particular interest in FFF. Usually, separations in FFF are influenced (but not directed) by surface properties of the sample components (to avoid particle-particle or particle-separator interactions). These properties can be modulated by the use of appropriate carrier-phase modifiers (surfactants). In terms of FFF separations, mass, size and density appear to be the major first order parameters. However, size is generally defined by the radius or the diameter of a sphere whose volume is identical to that of the cell. Size can therefore be deduced accurately if the cell of interest is perfectly spherical. However, this is not usually the case, and the sphericity index, I, is then used:

In this equation V is the cell volume and S is its surface area can be difficult to determine. In terms of cell population, these dimensions are averages and should be associated with a variance. These general considerations are also valid for cell density. Each measurable cell characteristic is therefore associated with an average value and its related variance. A probe of this 'diversity' is of great significance, even in a highly purified cell population. Polydispersity is historically defined in polymer chemistry as the ratio of the percentage of the standard deviation to the average value.

Any cell population can therefore be described by a set of values (average, variance) that leads to a polydispersity index. Cells are different from polymers in that each measurable characteristic of a cell can be associated with polydispersity, so polydispersity can apply to mass, size, volume or density. This matrix of parameters (average, variance) can be used to define a 'multipolydispersity matrix'. The dimensions of the multipolydispersity index depend on the information available. For example, the human normal red blood cell (HNRBC) is the best known cellular material. Its average volume has been measured as 95 + 5 fL, and its surface area has been calculated as 138.0 $ 5 |im2. The HNRBC is known for its discoid shape, which generates other measurable dimensions. Its average diameter is 8.1 + 0.43 |im, and minimum and maximum thicknesses of 1.0 + 0.3 |im and 2.4 + 0.15 |im have been measured. An average sphericity index of 0.77 has been calculated. The density was found to vary from 1.035 to 1.102, with an average value of 1.051. Using these data, it is possible to describe the HNRBC population using a preliminary multipolydispersity matrix, as shown in Table 1.

More complex properties such as cell/nucleus ratio or surface electric charge density (£ potential, used for dielectric cell purification) can later be added to this basic matrix. In FFF, the sample (i.e. cell surface) characteristics are most important in the development of any separation process. The separator material is chosen so as to avoid or limit particle-separator interactions, which can lead to limited recovery and viability, separator ageing or poisoning. In this domain a general set of empirical rules emerged. If the cell surface is hydrophilic (blood and yeast cells) a hydro-phobic material should be used for the separator. The lower the surface energy of the cell, the less nonspecific cell-separator interactions occurred, and vice

Table 1 Multipolydispersity matrix for HNRBC

Average

Standard deviation

Polydispersity

(%)

Volume (fL)

95

5

10

Density

1.051

0.08

3

Sphericity

0.77

0.15

11

versa. The interactions between the environmental material and the cells can now be assessed by the general 'biocompatibility' rules.

However the HNRBC population found in circulating blood is not a homogenous, and contains red blood cells (RBCs) of different ages. This suggests that a morphologically homogenous population can be considered to contain different sub-populations. An HNRBC suspension contains cells of equal volume, and cells of equal density, shape or mass. This complex definition of 'population' characteristics is described in Figure 1A. If a set of common parameters is defined, the HNRBCs that meet these limitations may represent only a minor proportion of the whole population. By considering some aspects of this multipolydispersity matrix, rules for cell separation based on biophysical characteristics can be found. Figure 1B shows a three-dimensional graphical representation of the multipolydispersity of the circulating blood cells (platelets, HNRBC, lymphocytes, monocytes, granulocytes) based on the characteristics of size and density. It is obvious that if a size driven cell separation technology could be used, all these populations could be isolated. An isolation technique based only on density would be more complex, as some blood cell population densities overlap. However a size and density driven separation (whose balance has not so far been assessed) would allow selective isolation.

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Solar Panel Basics

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