Abstract：
:Glycosylation of the Fc domain is an important driver of antibody effector function. While assessment of antibody glycoform compositions observed across total plasma IgG has identified differences associated with a variety of clinical conditions, in many cases it is the glycosylation state of only antibodies against a specific antigen or set of antigens that may be of interest, for example, in defining the potential effector function of antibodies produced during disease or after vaccination. Historically, glycoprofiling such antigen-specific antibodies in clinical samples has been challenging due to their low prevalence, the high sample requirement for most methods of glycan determination, and the lack of high-throughput purification methods. New methods of glycoprofiling with lower sample requirements and higher throughput have motivated the development of microscale and automatable methods for purification of antigen-specific antibodies from polyclonal sources such as clinical serum samples. In this work, we present a robot-compatible 96-well plate-based method for purification of antigen-specific antibodies, suitable for such population level glycosylation screening. We demonstrate the utility of this method across multiple antibody sources, using both purified plasma IgG and plasma, and across multiple different antigen types, with enrichment factors greater than 1000-fold observed. Using an on-column IdeS protease treatment, we further describe staged release of Fc and Fab domains, allowing for glycoprofiling of each domain.
journal_name
J Immunol Methodsjournal_title
Journal of immunological methodsauthors
Brown EP,Normandin E,Osei-Owusu NY,Mahan AE,Chan YN,Lai JI,Vaccari M,Rao M,Franchini G,Alter G,Ackerman MEdoi
10.1016/j.jim.2015.06.005subject
Has Abstractpub_date
2015-10-01 00:00:00pages
27-36eissn
0022-1759issn
1872-7905pii
S0022-1759(15)30007-7journal_volume
425pub_type
杂志文章abstract：:Fractionation of lactoperoxidase iodinated cell surface material on miniature DEAE-cellulose columns provided a rapid method for separating labeled lipid from cell surface antigens. The procedure also removed poorly solubilized aggregates yielding a labeled preparation which demonstrated stable, reproducible immunopre...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(77)90029-1
更新日期：1977-01-01 00:00:00
abstract：:Immunological techniques have been used to identify and quantitate rabbit antibody light chains in the urine of rabbits. The influence of these light chains on the radioallergosorbent test (RAST) used for the diagnosis of laboratory animal allergy to rabbit urinary protein allergens is described. The presence of IgG a...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(83)90009-1
更新日期：1983-06-24 00:00:00
abstract：:The antibody response in rabbits and chickens (serum and egg yolk) immunized with human immunoglobulin G was studied during a 1-year immunisation period comparing the efficacy of three adjuvants: Freund's complete adjuvant (FCA), Freund's incomplete adjuvant (FIA) and Hunter's TiterMax adjuvant (HTM). Monthly boosters...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(96)00010-5
更新日期：1996-05-27 00:00:00
abstract：:Frequencies of lipopolysaccharide (LPS)-reactive B cells in the mouse can be determined in the limiting dilution culture system developed by Andersson et al. (1976, 1977a) which is completely dependent upon the presence of thymus filler cells, usually of rat origin. The assessment of B cell clones of mouse origin, how...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(82)90399-4
更新日期：1982-01-01 00:00:00
abstract：:Immunoglobulins of the IgE and IgG classes have been causally associated with hypersensitivity reactions in man and in numerous animal species including mice, rats and guinea pigs. The use of the guinea pig as an animal model for both pulmonary and dermal hypersensitivity reactions, and the recent recognition of the i...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(91)90359-n
更新日期：1991-05-17 00:00:00
abstract：:Bacillus anthracis spores and Escherichia coli were stained with fluorescein-conjugated antibody using direct and indirect methods, then analyzed by means of a commercial flow cytometer. To reduce the cytometer's fluorescence component resulting from unreacted conjugate, reaction mixtures were either diluted or were c...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(87)90153-0
更新日期：1987-08-03 00:00:00
abstract：:Dengue virus (DENV) has substantial global impact, with an estimated 390million people infected each year. In spite of this, there is currently no approved DENV-specific vaccine or antiviral. One reason for this is the difficulty involved with development of an adequate animal model. While non-human primates support v...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2014.01.001
更新日期：2014-08-01 00:00:00
abstract：:As a tetrameric major histocompatibility complex (MHC) class I-peptide complex (tetramer) is capable of detecting antigen-specific cytotoxic T lymphocyte (CTL) by flow cytometry, significant information about the generation of in vivo immunity can be obtained. It is, however, difficult to make a soluble wild type of M...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(02)00329-0
更新日期：2002-12-20 00:00:00
abstract：:A sensitive method for staining proteins after transfer from polyacrylamide gels to nitrocellulose paper is described. Transferred proteins are first derivatized by reaction of the nitrocellulose replica with sulfosuccinimidobiotin and are then reacted sequentially with streptavidin, rabbit anti-streptavidin, and hors...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(86)90504-1
更新日期：1986-08-21 00:00:00
abstract：:The effect of various physical or chemical treatments of splenic stimulator cells on their endogenous, mitogen-inducible IL-2 production and on their ability to induce IL-2 production in clonally developing helper T lymphocytes was investigated. While most methods (T cell depletion by monoclonal antibodies plus comple...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(88)90241-4
更新日期：1988-05-09 00:00:00
abstract：:We have developed a large scale (10 g/batch) non-chromatographic method for the collection of IgM directly from plasma obtained from therapeutic plasmapheresis of patients. The technique uses sequential precipitations from ammonium sulfate and polyethylene glycol, and gives high yields of undenatured IgM antibodies. T...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(93)90252-3
更新日期：1993-01-14 00:00:00
abstract：:The measurement of the absolute CD4 T-cell count is critical in the initial evaluation and staging of HIV-infected persons, yet access to this technology remains limited in many low resource settings where disease burden is highest. Here we evaluate the performance of a prototype point-of-care device (POC) to quantify...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2012.10.002
更新日期：2013-01-31 00:00:00
abstract：:Irradiated human peripheral blood lymphoid cells undergo apoptosis and progressively exhibit typical changes in light scatter and plasma membrane integrity that can be easily tracked by flow cytometry. Using this model, we assessed the capacity of a newly developed fluorochrome, Apostain, in identifying early apoptosi...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(97)00054-9
更新日期：1997-06-23 00:00:00
abstract：:A method is described for determining the percentage of rosettes formed by sheep erythrocytes and human peripheral lymphocytes (ER) using cytofluorographic analysis (CFGA). The technique utilizes acridine orange dye for differentially staining nucleus and cytoplasm of the lymphocytes to distinguish them from erythrocy...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(76)90091-0
更新日期：1976-01-01 00:00:00
abstract：:Using in vitro established tumour cell lines attempts were made to assess the suitability of tetrazolium salt reduction (MTT) assay to replace the conventional radioactive base techniques for measuring cell proliferation and cell killing. The optimum conditions for MTT loading time, concentration of MTT and the time f...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(93)90012-v
更新日期：1993-03-15 00:00:00
abstract：:Rhesus monkey serum was applied to a protein A-Sepharose column at pH 7.3. IgG was not detectable in the unbound effluent. Bound protein was eluted with a pH gradient of citrate-phosphate buffer. Two major elution peaks reproducibly resulted at pH 4.85-4.9 and pH 4.35-4.43, respectively, with only slight variability a...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(82)90170-3
更新日期：1982-01-01 00:00:00
abstract：:Murine B16 melanoma cells and metastasis variants of this tumor are resistant to NK activity mediated by normal splenocytes. The B16 cells are, however, sensitive to splenocyte-mediated cytostasis. Cytostasis was measured by a [125I]UDR incorporation-inhibition [(125I]UDR I-I) assay. The main characteristics of the [1...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(81)90066-1
更新日期：1981-01-01 00:00:00
abstract：:A competitive radioimmunoassay for immunoconglutinin (IK) was developed using isolated antostimulated rabbit IK labelled with 125I and glutaraldehyde-fixed sheep erythrocytes coated with fixed complement (EAC). IKs in test sera of human or animal origin produced specific inhibition of uptake of [125I]IK onto EAC and t...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(80)90045-9
更新日期：1980-01-01 00:00:00
abstract：:The aim of the present study was to analyze the relations between the serum anti-erythropoietin antibody (AEAb) levels and the antibodies' neutralizing activity in 20 patients with renal anemia and rhEPO-induced antibodies. AEAb levels were determined by the enzyme-linked immunosorbent assay (ELISA, double antigen-bri...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2008.04.009
更新日期：2008-07-31 00:00:00
abstract：:Phycobilisomes are cyanobacterial photosynthetic energy transfer complexes partly composed of phycobiliproteins, proteins that are widely used as conjugable fluorochromes for flow cytometry. The brightness and photostability of phycobiliproteins suggest that intact phycobilisomes could constitute even brighter probes ...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/s0022-1759(01)00367-2
更新日期：2001-08-01 00:00:00
abstract：:Schistosoma japonicum glutathione S-transferase (GST), expressed from a pGEX plasmid, was isolated from Escherichia coli cells and used to immunize mice in order to generate specific anti-GST monoclonal antibodies. Using a modified immunization and fusion procedure, one stable hybridoma clone secreting an anti-GST ant...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(95)00204-9
更新日期：1995-12-15 00:00:00
abstract：:A method that permits high energy beta-irradiation to be applied to individual lymphocyte microplate cultures is described. The principle involves the introduction of single energy sources (in this case 32P) contained within tubes which rest inside wells containing the lymphocyte cultures. Cell viability and mitogen-i...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(84)90205-9
更新日期：1984-06-08 00:00:00
abstract：:Macrophage cell lines have been derived from cells residing in a number of tissues from a variety of species. In this report, the methods used to obtain these cell lines are reviewed and a simple and efficient method for generating nonvirus-transformed lines from individual cloned progenitors located in mouse tissues ...
journal_title：Journal of immunological methods
pub_type： 杂志文章,评审
doi：10.1016/0022-1759(94)90007-8
更新日期：1994-09-14 00:00:00
abstract：:A new system for assaying monoclonal antibodies consisting of an 8 x 12 array of sticks which fits into a 96-well microtiter plate is described. Tests using virus specific monoclonal antibodies and virus proteins demonstrated sensitivity equivalent to the conventional microtiter plate assay. Antibody production, antig...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(83)90356-3
更新日期：1983-03-25 00:00:00
abstract：:A rapid and simple immunostick immunosorbent ELISA has been developed for the diagnosis of myasthenia gravis. It is based on coating immunosticks with the major myasthenia gravis autoantigens acetylcholine receptor (AChR) or MuSK and it can be performed in <1 h, in a non-specialized examination place by non-skilled pe...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2018.06.016
更新日期：2018-09-01 00:00:00
abstract：:A simple method for the simultaneous isolation of IgG2, IgG1, IgA and IgM from porcine colostral whey is described. It makes use of DEAE-cellulose chromatography followed by repeated Sepharose 6B exclusion chromatography after the removal of fat, casein and lipoproteins, Fused rocket immunoelectrophoresis is used to s...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(77)90058-8
更新日期：1977-01-01 00:00:00
abstract：:There are several double immunolabelling methods but each has its drawbacks. More often than not, antibodies with the required specificities are available in only one species and their use normally produces false labels due to cross-reactivity. We describe a new and reliable technique for staining with primary antibod...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(95)00281-2
更新日期：1996-04-19 00:00:00
abstract：:A polyethylene glycol (PEG) radioimmunoprecipitation assay for human IgG is described that is sufficiently sensitive to detect 0.5 ng of IgG. This model antibody-antigen system was also used to study the stoichiometries of PEG-precipitated complexes. Our results suggest that the presence of PEG may affect the stoichio...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(79)90091-7
更新日期：1979-01-01 00:00:00
abstract：:Serum cytokine profiling is a powerful tool to link host immune defense with disease pathogenesis. Although several multiplex assays are commercially available, none has been rigorously validated in the context of chronic infectious disease (such as HIV infection). Here we compared the measurement of proinflammatory c...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/j.jim.2011.06.033
更新日期：2011-09-30 00:00:00
abstract：:An immunoaffinity method for the purification of hepatitis B surface antigen (HBsAg) using monoclonal anti-idiotype (anti-Id) as a specific eluent is described. Ammonium sulfate fractions of HBsAg-containing plasma were absorbed on monoclonal idiotype (Id) immunoadsorbents. The bound material was eluted from the colum...
journal_title：Journal of immunological methods
pub_type： 杂志文章
doi：10.1016/0022-1759(90)90019-r
更新日期：1990-01-24 00:00:00
