Methods for binding cells to plastic: Application to a solid-phase radioimmunoassay for cell-surface antigens

Stocker, John; Heusser, C H

doi:10.1016/0022-1759(79)90044-9

Cited by 134 publications

(25 citation statements)

References 9 publications

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“…This was essentially performed as previously described (38). Macrophages incubated overnight in RPMI 1640 medium containing penicillin (12.5 U/ml) and streptomycin (6.5 g/ml) were scraped into 15-ml polypropylene tubes and centrifuged at 400 rpm for 5 min at 4°C to remove broken cells.…”

Section: Detection Of Cell Surface Grp78 Using a Cellular Elisa (Celisa)mentioning

confidence: 99%

The Role of MTJ-1 in Cell Surface Translocation of GRP78, a Receptor for α2-Macroglobulin-Dependent Signaling

Misra

Gonzalez-Gronow

Gawdi

et al. 2005

The Journal of Immunology

111

105

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MTJ-1 associates with a glucose-regulated protein of Mr ∼78,000(GRP78) in the endoplasmic reticulum and modulates GRP78 activity as a chaperone. GRP78 also exists on the cell surface membrane, where it is associated with a number of functions. MHC class I Ags on the cell surface are complexed to GRP78. GRP78 also serves as the receptor for α2-macroglobulin-dependent signaling and for uptake of certain pathogenic viruses. The means by which GRP78, lacking a transmembrane domain, can fulfill such functions is unclear. In this study we have examined the question of whether MTJ-1, a transmembrane protein, is involved in the translocation of GRP78 to the cell surface. MTJ-1 and GRP78 coimmunoprecipitated from macrophage plasma membrane lysates. Silencing of MTJ-1 gene expression greatly reduced MTJ-1 mRNA and protein levels, but also abolished cell surface localization of GRP78. Consequently, binding of the activated and receptor-recognized form of α2-macroglobulin to macrophages was greatly reduced, and activated and receptor-recognized form of α2-macroglobulin-induced calcium signaling was abolished in these cells. In conclusion, we show that in addition to assisting the chaperone GRP78 in protein quality control in the endoplasmic reticulum, MTJ-1 is essential for transport of GRP78 to the cell surface, which serves a number of functions in immune regulation and signal transduction.

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Section: Detection Of Cell Surface Grp78 Using a Cellular Elisa (Celisa)mentioning

confidence: 99%

The Role of MTJ-1 in Cell Surface Translocation of GRP78, a Receptor for α2-Macroglobulin-Dependent Signaling

Misra

Gonzalez-Gronow

Gawdi

et al. 2005

The Journal of Immunology

111

105

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“…Ninety-six-well microtiter plates (Vangard, Neptune, NJ; NuncImmunoplate) were coated with poly (L-lysine) (21); live leishmanias (108 cells per ml in PJ/NaCl) were added to 50 Al per well. The plate was incubated at 4°C for 60 min to allow the cells to settle and then was submerged gently into Pi/NaCl containing glutaraldehyde (0.5%; Polysciences, Warrington, PA) for 15 min (22). After washings in Pi/NaCl containing 0.25% Tween 20 and 0.02% azide, the plates were stored in Pi/NaCl containing 0.5% gelatin and azide.…”

Section: Methodsmentioning

confidence: 99%

Surface antigenic change during differentiation of a parasitic protozoan, Leishmania mexicana: Identification by monoclonal antibodies.

Fong¹,

Chang²

1982

Proc. Natl. Acad. Sci. U.S.A.

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The fusion of SP2/O myeloma cells with spleen cells from mice immunized with Leishmania msexcana amazonensw promastigotes produced hybridoma clones. Indirect immunofluorescent antibody assay with live leishmanias showed that the monoclonal antibody 6H12 recognized only the antigens bound to the surface of L. mexicana amazonensus promastigotes. It also showed that the antibody bound to neither amastigotes ofthis species nor to other Leishmania species-i.e., L. braziliensis braziliensis, L. tropica, and L. donovani. Monoclonal antibodies from three other clones (4D11, 4H9, and 6A11) were found to compete with 6H12 for binding to L. mexicana promastigotes. With lysates of [asS]methionine-labeled promastigotes, all four monoclonal antibodies precipitated the same triplet set of protein bands at the -68,000-dalton region, whereas another monoclonal antibody (6G5) precipitated a different band at -90,000 daltons. During differentiation of L. mexicana amazonensis from amastigotes to promastigotes, there was a 4-to 8-fold increase above the initial level in the binding of 6H12 monoclonal antibody to leishmanias, as detected by enzyme-linked immunosorbent assay and quantitative fluorometric assay, respectively. Thus, we have demonstrated the use ofmonoclonal antibodies as probes for antigens that change during leishmanial differentiation.

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“…The cells were recovered after 3 or 4 days of culture, washed with PBS buffer, resuspended in the same buffer at a concentration of 1.5ϫ10 6 cells per milliliter, and immobilized on microtitration plates by using an adaptation of the method of Stocker and Heusser. 45 We introduced the following modifications: flexible polyvinyl chloride plates were first treated with polyglutaraldehyde during 2 hours at 22°C, excess glutaraldehyde was eliminated by washing the plates with distilled water, and 80 000 cells were added into each well. After centrifugation at 233g during 10 minutes, the cells were incubated in the plates for 1 hour at 22°C, followed by an incubation of 10 minutes in a bath of 0.25% polyglutaraldehyde; after 2 final washes with PBS, 100 L per well of PBS containing 4 mg/mL BSA and 0.1% thimerosal was added, and the plates were sealed and stored at 4°C until further use.…”

Section: Cell Culture and Preparation Of Cell Surfacesmentioning

confidence: 99%

Effect of Individual Plasma Lipoprotein(a) Variations In Vivo on Its Competition With Plasminogen for Fibrin and Cell Binding

Soulat

Loyau

Baudouin

et al. 2000

ATVB

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Abstract-Simultaneous natural changes in lipoprotein(a) [Lp(a)] and plasminogen occur in the nephrotic syndrome and offer a unique opportunity to investigate their effects on plasminogen activation under conditions fashioned in vivo. Plasminogen, Lp(a), and apolipoprotein(a) in plasma were characterized, and their competitive binding to carboxyterminal lysine residues of fibrin and cell membrane proteins was determined in nephrotic children during a flare-up of the disease (61 cases) and after 6 weeks (33 cases) and 6 months (42 cases) of remission. Low plasminogen concentrations (median 1.34 mol/L, range 0.39 to 1.96 mol/L) and high Lp(a) levels (median 0.27 g/L, range 0.07 to 2.57 g/L) were detected at flare-up. These changes were associated with an increased Lp(a) binding ratio onto fibrin (3.13Ϯ0.48) and cells (1.53Ϯ0.24) compared with binding ratios of control children (1.31Ϯ0.19 and 1.05Ϯ0.07, respectively) with normal plasminogen and low Lp(a) (median 0.071 g/L). After 6 weeks and 6 months of remission, the values for net decrease in Lp(a) binding to fibrin were 1.7Ϯ0.22 (after 6 weeks) and 1.88Ϯ0.38 (after 6 months) and were correlated with low Lp(a) concentrations (median 0.2 g/L, range 0.07 to 0.8 g/L; and median 0.12 g/L, range 0.07 to 1.34 g/L) and inversely associated with increased plasminogen levels (median 1.82 mol/L, range 1.4 to 2.1 mol/L; and median 1.58 mol/L, range 1.1 to 2.1 mol/L). These studies provide the first quantitative evidence that binding of Lp(a) to lysine residues of fibrin and cell surfaces is directly related to circulating levels of both plasminogen and Lp(a) and that these glycoproteins may interact as competitive ligands for these biological surfaces in vivo. This mechanism may be of relevance to the atherothrombotic role of Lp(a), particularly in nephrotic patients.

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Methods for binding cells to plastic: Application to a solid-phase radioimmunoassay for cell-surface antigens

Cited by 134 publications

References 9 publications

The Role of MTJ-1 in Cell Surface Translocation of GRP78, a Receptor for α2-Macroglobulin-Dependent Signaling

The Role of MTJ-1 in Cell Surface Translocation of GRP78, a Receptor for α2-Macroglobulin-Dependent Signaling

Surface antigenic change during differentiation of a parasitic protozoan, Leishmania mexicana: Identification by monoclonal antibodies.

Effect of Individual Plasma Lipoprotein(a) Variations In Vivo on Its Competition With Plasminogen for Fibrin and Cell Binding

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