Immunology

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    Protocols in Current Issue
    Protocol for High Throughput Screening of Antibody Phage Libraries
    [Abstract]

    Phage display is a proven and widely used technology for selecting specific antibodies against desired targets. However, an immense amount of effort is required to identify and screen the desired positive clones from large and diverse combinatorial libraries. On the other hand, the selection of positive binding clones

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    Identification of SARS-CoV-2 Neutralizing Antibody with Pseudotyped Virus-based Test on HEK-293T hACE2 Cells
    [Abstract]

    Neutralizing antibodies (NAbs) are of particular importance because they can prevent binding of the receptor binding domain (RBD) of the spike protein (S protein) to the angiotensin-converting enzyme 2 (ACE2) receptor present at the surface of human cells, preventing virus entry into the host cells. The gold standard method for detection of

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    A High-throughput Automated ELISA Assay for Detection of IgG Antibodies to the SARS-CoV-2 Spike Protein
    [Abstract]

    The SARS-CoV-2 pandemic and vaccination campaign has illustrated the need for high throughput serological assays to quantitatively measure antibody levels. Here, we present a protocol for a high-throughput colorimetric ELISA assay to detect IgG antibodies against the SARS-CoV-2 spike protein. The assay robustly distinguishes positive from negative

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    Production of Recombinant Replication-defective Lentiviruses Bearing the SARS-CoV or SARS-CoV-2 Attachment Spike Glycoprotein and Their Application in Receptor Tropism and Neutralisation Assays
    Authors:  Nazia Thakur, Giulia Gallo, Ahmed M. E. Elreafey and Dalan Bailey, date: 11/05/2021, view: 1985, Q&A: 0
    [Abstract]

    For enveloped viruses, such as SARS-CoV-2, transmission relies on the binding of viral glycoproteins to cellular receptors. Conventionally, this process is recapitulated in the lab by infection of cells with isolated live virus. However, such studies can be restricted due to the availability of high quantities of replication-competent virus,

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    Plasmodium cynomolgi Berok Growth Inhibition Assay by Thiol-reactive Probe Based Flow Cytometric Measurement
    Authors:  Jessica Jie Ying Ong, Bruce Russell and Jin-Hee Han, date: 09/05/2021, view: 1860, Q&A: 0
    [Abstract]

    The relapsing malaria species, Plasmodium vivax, is the most widely distributed and difficult-to-treat cause of human malaria. The merozoites of P. vivax preferentially invade ephemeral human CD71+ reticulocytes (nascent reticulocytes), thereby limiting the development of a robust continuous culture in vitro. Fortunately, P. vivax’s sister

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    Construction of a Highly Diverse mRNA Library for in vitro Selection of Monobodies
    [Abstract]

    Recently, we developed transcription/translation coupled with the association of puromycin linker (TRAP) display as a quick in vitro selection method to obtain antibody-like proteins. For the in vitro selection, it is important to prepare mRNA libraries among which the diversity is high. Here, we describe a method for the preparation of monobody

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    Production of the Receptor-binding Domain of the Viral Spike Proteins from 2003 and 2019 SARS CoVs and the Four Common Human Coronaviruses for Serologic Assays and Inhibitor Screening
    [Abstract]

    The recombinant receptor-binding domain (RBD) of the viral spike protein from SARS-CoV-1 and 2 are reliable antigens for detecting viral-specific antibodies in humans. We and others have shown that the levels of RBD-binding antibodies and SARS-CoV-2 neutralizing antibodies in patients are correlated. Here, we report the expression and purification

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    An Image-based Dynamic High-throughput Analysis of Adherent Cell Migration
    [Abstract]

    In this protocol, we describe a method to monitor cell migration by live-cell imaging of adherent cells. Scratching assay is a common method to investigate cell migration or wound healing capacity. However, achieving homogenous scratching, finding the optimal time window for end-point analysis and performing an objective image analysis imply, even

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    Multiplication and Growth Inhibition Activity Assays for the Zoonotic Malaria Parasite, Plasmodium knowlesi
    Authors:  Franziska Mohring, Thomas A. Rawlinson, Simon J. Draper and Robert W. Moon, date: 09/05/2020, view: 3617, Q&A: 0
    [Abstract] Malaria remains a major cause of morbidity and mortality globally. Clinical symptoms of the disease arise from the growth and multiplication of Plasmodium parasites within the blood of the host. Thus in vitro assays to determine how drug, antibody and genetic perturbations affect the growth rate of Plasmodium parasites ...
    Assessments of HLA-I Specificities of Anti-HLA-I Monoclonal Antibodies Using Solid Phase Bead Arrays
    Authors:  Anita J. Zaitouna, Daniel S. Ramon and Malini Raghavan, date: 06/20/2020, view: 3341, Q&A: 0
    [Abstract] Human leukocyte antigen class I (HLA-I) molecules are a group of structurally-related cell surface proteins with a high degree of variability within the population. While only up to six variants are expressed in an individual person, the whole population contains thousands of different variants. The ability to distinguish specific variants is ...



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