**7. Capillary electrophoresis (CE)**

Jorgenson and Lukacs in 1981 invent capillary electrophoresis (CE) most often termed as capillary zone electrophoresis (CZE) [34]. It is a type of electrophoresis in which analytes are separated by applying an electric field across buffer solutionfilled capillary tubes. The proposed instrumental technique was later implemented in a number of applications such as bioanalytical [35] and forensic drug analysis [36]. This technique is an alternative to the gel electrophoresis or LC (**Figure 6**).

CZE method correlated well with an automated kinetic fluorescent assay. An example is analysis of NAG by CE after incubation of urine samples using synthetic substrate, methylumbelliferyl-β-d-glucosaminide [37].

**Figure 6.** *Capillary electrophoresis separation method.*

#### **8. Microscopy**

In the simplest microscopic methods, a specimen is illuminated by visible light and observed either against a bright background (bright-field microscopy) or a dark background (dark-field microscopy). The presence of cells that are not expected in the healthy person may be an indicator of disease. For example, a simple microscopic analysis of blood sample identifies the sickle cell anemia, and analysis of urine quantifies the presence of pus cells, which is an indicator of infection.

Light microscopy uses light as the illumination radiation. This is used to identify the microorganisms based on their morphology. An application of microscopy is to count the number of different cells per unit volume of blood or any other sample using a hemocytometer. Fluorescence microscopy has emerged as a very powerful tool for studying molecular processes owing largely to the advancement in optics and discovery of the green fluorescent protein and development of its analogs with different spectral properties [38]. Several advancements in the field of fluorescence microscopy have been achieved that includes the following techniques:

• Confocal laser scanning microscopy (CLSM)

CLSM is a type of fluorescence microscopy that allows imaging of the samples at different focal planes that light emitting from below or above the desired focal plane is eliminated. This results in very high lateral resolution and allows determining the spatial localization of the molecules [39].

• Total internal reflection fluorescence (TIRF) microscopy

TIFR is another type of fluorescence microscopy wherein the optics allows imaging of the molecules that are almost like microscopic slide. The resolution of light microscopes depends on the wavelength of the light used. The smaller the

**39**

*Advancement in Analytical and Bioanalytical Techniques as a Boon to Medical Sciences*

wavelength of the light used, the better the resolution obtained. Wavelength of the visible light imposes a resolution limit of ~0.2 μm on the light microscopes. Hemocytometer (Neubauer chamber) is a glass slide which has a counting chamber at the center. A glass cover is placed on the hemocytometer, and the sample is gently introduced into the chamber. The sample chamber has a grid which allows counting

Gene therapies are considerable improvements over the existing therapy because of the advantage in dosing schedule, patient compliance, toxicity, immunogenicity, and cost. Owing to this, gene therapy provides novel approaches for the treatment of inherited and acquired diseases. The development of a nonviral gene delivery vehicle capable of efficient, cell-specific delivery will be a valuable addition to the clinical armamentarium [41]. For example, the liver places a central role in the metabolism and production of serum proteins; it is an important target organ for gene therapy. Hepatic metabolic diseases as well as acquired diseases may also serve as targets for hepatic gene therapy [42]. Most recent gene therapy protocols describe

delivery of foreign genes by means of injecting lentiviral particles [43].

Immunoassays are the quantitation of bioanalyte that depends on the reaction of an antigen (analyte) and an antibody. These methods are based on a competitive binding reaction between a fixed amount of labeled form of an analyte and a variable amount of unlabeled sample analyte for a limited amount of binding sites on a highly specific anti-analyte antibody. When immunoanalytical reagents (analyte or antibody) are mixed and incubated, the analyte is bound to the antibody forming an immune complex. This complex is separated from the unbound reagent fraction by physical or chemical separation technique. Analysis is achieved by measuring the label activity (e.g., radiation, fluorescence, or enzyme) in either of the bound or free fraction [44]. Immunoassay methods have been widely used in many important areas such as diagnosis of diseases, therapeutic drug monitoring, clinical pharmacokinetic and bioequivalence studies in drug discovery, and pharmaceutical industries. A few

• Radioimmunoassay (RIA) methods have been used successfully for the determination of a limitless number of pharmaceutically important compounds in biological fluids. RIA is used to analyze thyroid hormone testing in patients

• Enzyme immunoassay (EIA) is analogous to RIA except that the label is an

• Chemiluminescence immunoassay (CLIA) involves a chemiluminescent

• Fluoroimmunoassay (FIA) is analogous to RIA except that the label is a fluoro-

• Liposome immunoassay (LIA) is the assay involving a liposome-encapsulating

**10. Immunological and radioisotope techniques**

immunoassays based on different labels are as follows:

after iodine-131 therapy.

substance as a label.

marker.

enzyme rather than a radioisotope.

phore rather than a radioisotope.

*DOI: http://dx.doi.org/10.5772/intechopen.80279*

**9. Gene therapy protocol**

of cells in a defined region using a microscope [40].

*Advancement in Analytical and Bioanalytical Techniques as a Boon to Medical Sciences DOI: http://dx.doi.org/10.5772/intechopen.80279*

wavelength of the light used, the better the resolution obtained. Wavelength of the visible light imposes a resolution limit of ~0.2 μm on the light microscopes. Hemocytometer (Neubauer chamber) is a glass slide which has a counting chamber at the center. A glass cover is placed on the hemocytometer, and the sample is gently introduced into the chamber. The sample chamber has a grid which allows counting of cells in a defined region using a microscope [40].
