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Antibodies and Antisera
Published in Lars-Inge Larsson, Immunocytochemistry: Theory and Practice, 2020
Another second reagent that can give rise to positive staining controls is avidin/streptavidin, employed in biotin-based techniques. Native egg-white avidin is a strongly basic molecule that shows unspecific binding to many tissues. Native avidin is therefore rarely used in immunocytochemistry today. Treatment of avidin with amino-group blockers reduces its negative charge and its unspecific bindings. The majority of avidin preparations available today have been treated in this way. Alternatively, streptavidin, having a neutral isoelectric point, can be employed (cf. Chapter 3, Section II.D). Even with these precautions, unspecific staining of tissues rich in endogenous biotin may occur. (Cells cultured in rich media are particularly troublesome.)
Immunohistochemistry of the Pulmonary Extracellular Matrix
Published in Joan Gil, Models of Lung Disease, 2020
Antonio Martinez-Hernandez, Peter S. Amenta
Another alternative is the avidin-biotin system, based on the high-affinity between avidin and biotin (Heitzmann and Richards, 1974). The antibody is labeled with biotin, followed by a avidin enzyme conjugate. Egg-white avidin is highly glycosylated, resulting in relatively high backgrounds (Heitzmann and Richards, 1974; Wooley and Longsworth, 1942). This problem can be overcome using strep-tavidin, a nonglycosylated protein of bacterial origin (Heitzmann and Richards, 1974; Wooley and Longsworth, 1942). In our hands, the strepavidin-biotin system seems more sensitive than the PAP system.
Experimental Stomatology
Published in Samuel Dreizen, Barnet M. Levy, Handbook of Experimental Stomatology, 2020
Samuel Dreizen, Barnet M. Levy
Due to the avidin content of the egg white, the stage was set for the development of an uncomplicated biotin deficiency. Avidin is a protein capable of binding with biotin rendering it nutritionally inactive. Although the patient had noticed mild dryness of the skin and occasional soreness of the mouth and lips, it was not until several months prior to the hospital admission that the symptoms increased in severity. The tongue became sore and reddened and the lips were fissured, encrusted, and bled occasionally. She had mild dysphagia, nausea, and vomiting. Associated with these symptoms were increasing lassitude, malaise, exertional dyspnea, easy fatigability, and substernal discomfort partially relieved by belching.
Synergic fabrication of pembrolizumab loaded doxorubicin incorporating microbubbles delivery for ultrasound contrast agents mediated anti-proliferation and apoptosis
Published in Drug Delivery, 2021
Huilin Liu, Xing Li, Zihe Chen, Lianjie Bai, Ying Wang, Weiyang Lv
Poly(lactic-co-glycolic acid) (PLGA; 50% lactide, 50% glycolide, MW = 10,000 Da) was purchased from Shandong Shuyuan Biotechnology Co., Ltd (Shandong, China). Poly(vinyl alcohol) (PVA, 87–89%, MW = 31,000–50,000) was obtained from Sigma-Aldrich (St Louis, MO). DOX was obtained from Shenzhen Wanle Pharmaceutical Co., Ltd. (Shenzhen, China). EZ-LinkTM Sulfo-LC-Biotinylation kit was purchased from Thermo Fisher Scientific, Inc. (Rockford, ILz. Amine-Peg2000-Biotin was purchased from Nanjing Ling Di Ren Chemical Technology Co., Ltd (Nanjing, China). Avidin and dyLight488-labeled avidin were obtained from Wuhan Boster Biotechnology Co., Ltd (Wuhan, China). Pembrolizumab was obtained from Hoffmann-La Roche, Inc (Little Falls, NJ). Cell Counting Kit-8, the Annexin V-FITC cell apoptosis detection kit, and the TUNEL apoptosis detection kit were purchased from Beyotime Biotechnology Co., Ltd (Shanghai, China). All chemicals were analytical grade and used without further processing.
Engineering of fluorescent or photoactive Trojan probes for detection and eradication of β-Amyloids
Published in Drug Delivery, 2020
Amal A. Aziz, Rafat A. Siddiqui, Zareen Amtul
IgG, as a fusion protein, cannot be engineered for certain biopharmaceuticals, such as oligopeptides, sequence-specific short interfering RNA, or antisense drugs (Pardridge, 2008). The in vivo delivery of such biotherapeutics or diagnostics through the BBB could be made possible by highly stable avidin-biotin bonding between the monobiotinylated fusion protein (biotin moieties are attached using –SS- or –XX- linkers) and the (streptavidin or neutral light) avidin-labeled TfRMAb (McMahon, 2008). Where SS may be a cleavable disulfide (thioether) 8-atom spacer and XX is a non-cleavable (amide) 14-atom spacer linker (McMahon, 2008). Due to a very strong (kD 10–15 M) affinity and dissociation (half-life 89-day) (Green, 1975) of avidin-biotin complex, the biotinylated fusion protein is instantaneously captured by the avidin-labeled TfRMAb (Boado et al., 2008). Placement of a biotin molecule at the 3′ end of the phosphodiester antisense oligodeoxynucleotides not only facilitates the attachment but also renders its degradation by the 3′-exonuclease (Asseline et al., 1984). Nearly any mono-biotinylated fusion protein or biopharmaceutical could be carried to the brain with the avidin-labeled TfRMAb that could serve as a universal Trojan horse. Additionally, avidin is shown to elicit no immunologic reactions when administered to humans (Samuel et al., 1996) (Figure 2(C)).
Combining the best of two worlds: highly flexible chimeric antigen receptor adaptor molecules (CAR-adaptors) for the recruitment of chimeric antigen receptor T cells
Published in mAbs, 2019
Diana Darowski, Sebastian Kobold, Christian Jost, Christian Klein
Avidin and streptavidin are both naturally occurring biotin-binding proteins with high affinities, but most of the monomeric or dimeric constructs reported so far are unstable, prone to aggregation, and reveal low biotin affinity. Several approaches have been undertaken to design monomeric or dimeric variants of avidin or streptavidin, respectively.69,70 Making use of modified avidin as ECD incorporated into a CAR, Urbanska et al. designed biotin-binding modCAR-T cells. The group showed that only modCAR-T cells expressing dimeric avidin were able to redirect biotinylated mAb- or scFv-CAR-adaptors. ModCAR-T cells expressing monomeric avidin did not show specific retargeting or activation of T cells. Since biotin is present in human plasma, the group demonstrated that, even at supra-physiological levels, soluble biotin did not cause antigen-independent activation of dimeric avidin engineered modCAR-T cells and did not inhibit the modCAR activity by acting as a competitor molecule.23