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Methods in molecular exercise physiology
Published in Adam P. Sharples, James P. Morton, Henning Wackerhage, Molecular Exercise Physiology, 2022
Adam P. Sharples, Daniel C. Turner, Stephen Roth, Robert A. Seaborne, Brendan Egan, Mark Viggars, Jonathan C. Jarvis, Daniel J. Owens, Jatin G. Burniston, Piotr P. Gorski, Claire E. Stewart
As discussed previously, both acute exercise stimuli and long-term exercise training can modulate changes in protein activity (phosphorylation), protein abundance and the location of a protein within skeletal muscle cells or fibres. Acutely, changes in activity, localisation or the co-localisation of two proteins may be indicative of ongoing signalling cascades, or protein interactions pertinent to skeletal muscle responses to exercise or nutritional intake (65, 66). Similarly, to Western blot, immunohistochemistry takes advantage of the binding of monoclonal or polyclonal antibodies that are complementary to the antigens on a target protein of interest. Monoclonal antibodies will only bind to one epitope on the antigen molecule on the protein of interest, whereas polyclonal antibodies will bind to multiple epitopes on the target antigen. Once bound, a secondary antibody conjugated to an enzymatic or fluorescent molecule can be added which will bind to the primary antibody. As immunohistochemistry is reliant on tissue being preserved as it where in-situ, using chemical fixation or snap-freezing rather than being homogenised, researchers are able to visualise the protein itself using miscroscopy.
Immunological Tests for Diagnosis of Disease and Identification of Molecules
Published in Julius P. Kreier, Infection, Resistance, and Immunity, 2022
The enzyme-linked immunosorbent assay (ELISA) technique is a highly versatile, sensitive, and quantitative procedure that requires little equipment and for which reagents are readily available. It was first introduced in 1971 by Engvall and Perlmann. The term enzyme immunoassay (EIA) is an alternative name for this type of test and is used for the many variants of the assay. Enzymes such as alkaline phosphatase or peroxidase can be linked to antibody without destroying either the antibody′s specificity or the enzyme′s activity. The enzyme acts as a label which makes detection of the antibody possible. Both monoclonal and polyclonal antibodies can be used in this type of test. Enzyme labels are cheaper, simpler to measure, and far more stable than radioactive labels. For these reasons, ELISA or EIA assays have in many cases replaced RIA and have done so while maintaining sensitivity and specificity. Many of the assays for hepatitis A and B in use today are based on ELISA. In addition to the use of enzymes in immunoassays, nonenzy-matic markers can also be used if they can be conjugated to the antibody (e.g., colloidal gold with silver enhancement, and biotination with avidine).
Peptide Vaccines in Cancers
Published in Mesut Karahan, Synthetic Peptide Vaccine Models, 2021
Öznur Özge Özcan, Rümeysa Rabia Kocatürk, Fadime Canbolat
Briefly, a variety of cancer immunotherapies have been developed for cancer treatments, including immune checkpoint blockage therapy, cytokine therapy, cancer vaccine, and chimeric antigen receptor (CAR)-T cell therapy and many have been studied (Riley et al. 2019). Monoclonal and polyclonal antibodies, antibodies, cytokines, cellular immunity, and vaccines used in cancer immunotherapy have become increasingly successful therapeutic agents in both preclinical models and clinical trials, particularly in the treatment of solid cancers. Histologically, these alternative methods of immunotherapy are used to reduce the size of solid cancer and in combination treatments with chemotherapy (Baxevanis et al. 2009). Another aim of tumor immunotherapy is to enable individuals with a high family history of cancer to gain immunity against target tumors, actively or passively (Kakimi et al. 2016). With the increasing advances in cancer immunotherapy, it has become possible to control both natural and adaptive immune cells (illustrated in Figure 11.1) and their responses on tumor cells (Zhang and Chen 2018).
Chicken egg yolk antibodies (IgY)-based antivenom for neutralization of snake venoms: a review
Published in Toxin Reviews, 2022
Ankit Choraria, Rajeswari Somasundaram, S. Janani, Selvakumar Rajendran, Naoual Oukkache, A. Michael
Monoclonal antibodies are considered to be one of the most promising tools in basic and applied research used for medical diagnosis and effective therapy. Due to its various applications in the field of drug discovery and high cost of production of mAbs using hybridoma technology, numerous researches have been carried out to find an alternative method for mAbs production. Among the other forms of antibodies, the antigen binding fragment (Fab) or the single chain fragment variable (scFv) displayed on phage is suitable and fast for specific antibodies selection. Chickens are considered to be an adequate animal model as they are most convenient and rapid host for constructing antibody library for selecting specific scFv against various targets (Lee et al. 2016a, 2016b). There have been reports for monoclonal IgY antibody production using hybridoma technology (Nakamura et al. 2003) but was not successful as IgY cannot bind to protein A or protein G. Chicken IgY antibodies are polyclonal in nature and, thus, have its own limitations for diagnosis purpose. Monoclonal IgY antibodies can be produced by genetic engineering technology (Zhang et al. 2010). Monoclonal antibody against one epitope will have lower efficacy than polyclonal antibodies against many epitopes. In case of snake toxins neutralization, a combination of various monoclonal antibodies has potential to neutralize snake venom and having ability to reduce symptoms from snake bite, thereby preventing death of envenomated patients (Lee et al. 2017a, 2017b).
Functionalized liposomes as drug nanocarriers for active targeted cancer therapy: a systematic review
Published in Journal of Liposome Research, 2022
Hanieh Abbasi, Nadereh Rahbar, Maryam Kouchak, Parna Khalil Dezfuli, Somayeh Handali
One of the important ways to treat cancer is to use formulations that deal directly or indirectly with the immune system. Today, antibody-antigen reactions are widely used to diagnose and treat many diseases. mAbs are antibodies produced by B cells. In contrast to polyclonal antibodies, mAbs are mono-specific and homogeneous (Breedveld 2000). Purified antibodies have more tendencies for binding to the antigens and as a result, the reactions are more sensitive. Until now various types of antibody fragments such as anti-CD20 monoclonal antibody, anti-CD47 monoclonal antibody, EGFR antibody, and anti-Fas monoclonal antibody have been used as targeting agents (Muhamad et al.2018). mAbs can strengthen the immune system's attack to cancer cells by binding to cancer cell surface antigens. The conjugation of antibodies to liposomes leads to improve the cancer cellular uptake and enhances the cytotoxic activity of the drugs (Waldmann 1991, Bayer 2019). All the reported researches in mAbs conjugated liposomes are summarized and given in Table 4 and Supplementary Table S3.
A comparative multi-tiered immunogenicity assessment of biosimilar pegylated filgrastim: validation of methods for clinical assessment of INTP5
Published in Expert Opinion on Biological Therapy, 2022
Pallavi Hajela, Ronak Patel, Prashant Kale, Manish Kumar, Sridevi Khambhampaty
Sensitivity of the assay is an important criteria for the acceptability in immunogenicity studies, and in general, sensitivities of below 100 ng/ mL are considered acceptable. However, the biggest challenge in immunogenicity assessment is the non-availability of human antibody controls. Surrogate controls, generally derived from a suitable animal species, are utilized to develop the assays. The assay must also be designed appropriately so that the assay format remains the same for the positive control (animal derived) to actual study sample (human anti-drug antibodies). The sensitivity of the method depends on the method design as well as on the positive control used. It is typical to screen various available polyclonal antibodies to select a suitable positive control. In the present publication, the positive controls were screened and selected for assuring that the assay can detect anti-pegfilgrastim, anti-filgrastim, and anti-peg antibodies in binding assays, and anti-filgrastim positive control was used for assessing the neutralizing bioactivity. The anti-pegfilgrastim was raised against the biosimilar product, while other antibodies were commercially sourced.