Immunohistochemistry Monoclonal Antibodies Development

Saura Airiza^*
*Correspondence: Saura Airiza, Department of Life Sciences, University of the Basque Country, Vitoria-Gasteiz, Spain, Email:

Keywords

Chromogens • Cells • Chemical • Histopathology

Introduction

In biological and medical research, chromogens are commonly used as substrates in enzymatic reactions that produce a colored product. Enzymes such as peroxidase, alkaline phosphatase, and β-galactosidase catalyze reactions that produce a chromogenic substrate into a colored product that can be detected and quantified. For example, 3,3'-diaminobenzidine (DAB) is a commonly used chromogen for immunohistochemical staining of tissue sections. When DAB is oxidized by peroxidase, it produces a brown precipitate that can be visualized under a microscope. Similarly, alkaline phosphatase substrates such as nitroblue tetrazolium and 5-bromo-4-chloro-3-indolyl phosphate produce blue and purple precipitates, respectively, when they are dephosphorylated by the enzyme.

Chromogens can also be used in colorimetric assays to detect and quantify the concentration of a specific molecule in a sample. For example, the Bradford assay is a widely used method for protein quantification that uses the chromogen Coomassie Brilliant Blue. When the dye binds to proteins in a sample, it undergoes a shift in its absorption spectrum that results in a color change from brown to blue. The intensity of the blue color is proportional to the protein concentration in the sample and can be measured spectrophotometrically [1].

Literature Review

In addition to their use in biological and chemical assays, chromogens are also used in the textile industry as dyes. Synthetic chromogens such as azo dyes, anthraquinone dyes, and phthalocyanine dyes are commonly used to dye textiles due to their bright colors, high colorfastness, and versatility. Azo dyes, for example, are widely used because they can be easily synthesized in a wide range of colors and shades, and they have good colorfastness to washing and light. However, the use of synthetic chromogens in the textile industry has raised concerns about their potential environmental and health impacts. Azo dyes, in particular, have been found to be toxic to aquatic life and can cause allergic reactions in humans. As a result, there has been growing interest in the development of natural chromogens as alternatives to synthetic dyes. Natural chromogens such as indigo, henna, and cochineal have been used for centuries to dye textiles and are generally considered to be safe and environmentally friendly.

Discussion

In recent years, there has been increasing interest in the use of chromogens in nanotechnology and material science. Nanoparticles coated with chromogenic molecules have been developed for use in sensors and electronic devices that can detect changes in temperature, humidity, and other environmental factors. For example, gold nanoparticles coated with chromogenic molecules such as thiolated 4-nitrobenzenethiol can be used as colorimetric sensors for detecting trace amounts of mercury in water. When the mercury binds to the thiol group on the nanoparticle surface, it causes a shift in the absorption spectrum of the chromogen, resulting in a color change that can be easily detected.

Chromogens are chemical compounds that are widely used in various fields, including biology, chemistry, and medicine. Chromogens are molecules that undergo chemical reactions with other compounds to produce colored products. The color change can be detected visually, and the intensity of the color can be measured using spectrophotometers. Chromogens are used as detection reagents in various assays, including immunohistochemistry, immunofluorescence, enzyme-linked immunosorbent assays (ELISA), and many others. In this article, we will explore the different types of chromogens, their mechanisms of action, and their applications [2].

DAB is a commonly used chromogen in immunohistochemistry assays. DAB reacts with peroxidase enzymes to produce a brown color. DAB is often used in conjunction with hydrogen peroxide, which acts as a catalyst to enhance the reaction. DAB is stable and produces a permanent brown stain, which is ideal for histological staining. AEC (3-Amino-9-ethylcarbazole) is another commonly used chromogen in immunohistochemistry assays. AEC produces a red color when it reacts with peroxidase enzymes. AEC is more sensitive than DAB and produces a brighter color. However, AEC is less stable than DAB and fades over time. Fast Red is a chromogen that is often used in immunohistochemistry assays. Fast Red produces a bright red color when it reacts with alkaline phosphatase enzymes. Fast Red is stable and produces a permanent stain. BCIP (5-Bromo-4-chloro- 3-indolyl-phosphate) and NBT (Nitro blue tetrazolium) are chromogens that are commonly used in conjunction with alkaline phosphatase enzymes. BCIP/NBT produces a blue-purple color when it reacts with alkaline phosphatase enzymes. BCIP/NBT is stable and produces a permanent stain [3,4].

Chromogens undergo chemical reactions with other compounds to produce colored products. The mechanism of action of chromogens varies depending on the type of chromogen and the enzyme or molecule it is reacting with. In the case of DAB, the chromogen reacts with peroxidase enzymes to produce a brown color. Peroxidase enzymes are often used as detection reagents in immunohistochemistry assays to detect the presence of a specific protein. When a tissue section is incubated with a primary antibody that recognizes the target protein, the antibody binds to the protein, forming an antibody-protein complex. A secondary antibody that is conjugated to peroxidase enzyme is then added, which binds to the primary antibody-protein complex. The peroxidase enzyme then reacts with hydrogen peroxide, which is also added to the tissue section, to produce a brown color [5,6].

Conclusion

In the case of Fast Red, the chromogen reacts with alkaline phosphatase enzymes to produce a bright red color. Alkaline phosphatase enzymes are often used as detection reagents in immunohistochemistry assays to detect the presence of a specific protein. When a tissue section is incubated with a primary antibody that recognizes the target protein, the antibody binds to the protein, forming an antibody-protein complex. A secondary antibody that is conjugated to alkaline phosphatase enzyme is then added, which binds to the primary antibodyprotein complex. The alkaline phosphatase enzyme then reacts with Fast Red to produce a bright red color.

Acknowledgement

None.

Conflict of Interest

There are no conflicts of interest by author.

References

1. Kato, Motohiro and Atsushi Manabe. "Treatment and biology of pediatric acute lymphoblastic leukemia." Pediatr Int 60 (2018): 4-12.

Google Scholar, Crossref, Indexed at

2. Subramaniam, Priya, K. Girish Babu and J. Nagarathna. "Oral manifestations in acute lymphoblastic leukemic children under chemotherapy." J Clin Pediatr Dent 32 (2008): 319-324.

Google Scholar, Crossref, Indexed at

3. Donnelly, J. Peter, Lisa A. Bellm, Joel B. Epstein and Stephen T. Sonis, et al. "Antimicrobial therapy to prevent or treat oral mucositis." Lancet Infect Dis 3 (2003): 405-412.

Google Scholar, Crossref, Indexed at

4. Sonis, Stephen T., Linda S. Elting, Dorothy Keefe and Douglas E. Peterson, et al. "Perspectives on cancer therapy‐induced mucosal injury: Pathogenesis, measurement, epidemiology, and consequences for patients." Int J Cancer 100 (2004): 1995-2025.

Google Scholar, Crossref, Indexed at

5. Navone, Roberto, Monica Pentenero and Sergio Gandolfo. "Liquid-based cytology in oral cavity squamous cell cancer." Curr Opin Otolaryngol Head Neck Surg 19 (2011): 77-81.

Google Scholar, Crossref, Indexed at

6. Brenner, Hermann, Peter Kaatsch, Tatjana Burkhardt‐Hammer and Dörthe O. Harms, et al. "Long‐term survival of children with leukemia achieved by the end of the second millennium." Cancer: Interdisciplinary International Journal of the American Cancer Society 92 (2001): 1977-1983.

Google Scholar, Crossref, Indexed at