The labelling of proteins to high specific radioactivities by conjugation to a 125I-containing acylating agent. Application to the radioimmunoassay

Bolton, A.E.; Hunter, W. M.

doi:10.1042/bj1330529

Cited by 3,217 publications

(1,030 citation statements)

References 19 publications

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“…'251-Labelling of proteins followed [22] or was by lactoperoxidase catalysis as in [20]. The average iodine binding was between 0.04-0.06 mol 1251/mol C2.…”

Section: Methodsmentioning

confidence: 99%

Human complement subcomponent C2: purification and proteolytic cleavage in fluid phase by C1̄s, C1̄r₂–C1̄s₂ AND 1̄

et al. 1982

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“…'251-Labelling of proteins followed [22] or was by lactoperoxidase catalysis as in [20]. The average iodine binding was between 0.04-0.06 mol 1251/mol C2.…”

Section: Methodsmentioning

confidence: 99%

Human complement subcomponent C2: purification and proteolytic cleavage in fluid phase by C1̄s, C1̄r₂–C1̄s₂ AND 1̄

et al. 1982

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“…IZ5I-labeled mIL-6 was prepared using the chloramine T procedure (Greenwood et al, 1963), and 12'I-labeled hlL-6 was prepared using the Bolton and Hunter procedure (Bolton & Hunter, 1973). Radiolabeled protein was separated from free iodine as described previously (Nicola & Metcalf, 1984;Ward & Simpson, 1991).…”

Section: '2si-iabeling Of Mil-6mentioning

confidence: 99%

Role of the C‐terminus in the activity, conformation, and stability of interleukin‐6

et al. 1993

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Two murine interleukin-6 (mIL-6) variants were constructed using the polymerase chain reaction (PCR), one lacking the last five residues (183-187) at the C-terminus (pMC5) and another with the last five residues of mIL-6 substituted by the corresponding residues of human IL-6 (pMC5H). The growth stimulatory activity of pMC5 on the mouse hybridoma cell line 7TD1 was <0.05% of mIL-6, whereas pMC5H and mIL-6 were equipotent. The loss of biological activity of pMCS correlated with its negligible receptor binding affinity on 7TD1 cells, while the binding of pMC5H was comparable to that of mIL-6. Both pMC5 and pMCSH, like mIL-6, failed to interact with recombinant soluble human IL-6 receptor when assayed by surface plasmon resonance-based biosensor analysis. These studies suggest that the C-terminal seven amino acids of human IL-6, alone, do not define species specificity for receptor binding.A variety of biophysical techniques, as well as the binding of a conformational-specific monoclonal antibody, indicated that the global fold of the mIL-6 variants was similar to that of mIL-6, although small changes in the NMR spectra, particularly for pMC5, were observed. Some of these changes involved residues widely separated in the primary structure. For instance, interactions involving Tyr-22 were influenced by the C-terminal amino acids suggesting that the N-and C-termini of mIL-6 are in close proximity. Equilibrium unfolding experiments indicated that pMC5 was 0.8 kcal/mol less stable than mIL-6, whereas pMCSH was 1.4 kcal/mol more stable. These studies emphasize the structural importance of the C-terminal amino acids of IL-6 and suggest that truncation or mutation of this region could lead to small but significant alterations in other regions of the molecule.

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“…ANTIBODY LABELLING (13,14) Bolton-Hunter reagent (0.5-1 mCi, 1700 Ci/mmol) supplied in benzene was dried by a gentle stream of nitrogen gas. Purified antiferritin IgG (45 p-g) was added in 50 p-L 0.1 mol/L sodium borate buffer, pH 8.5.…”

Section: Antibody Production and Purificationmentioning

confidence: 99%

Measurement of ferritin in serum: Application in Diagnostic use

Oost¹,

Beld²,

Cloin³

et al. 1984

Clinical Biochemistry

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A sandwich-type radioimmunoassay for serum ferritin was developed using iron-rich human liver ferritin and evaluated for its clinical usefulness. In young healthy males and females, the mean serum ferritin concentrations were 44 ~g/L (range 7-158) and 16 p.g/L (range 4-56), respectively. In anemic patients lower serum ferritin concentrations were found, while in most patients with iron overload serum ferritin concentrations well above 1000 ~g/L were measured. Comparison of our method with a commercially available radioimmunoassay kit revealed a good correlation, except for sera with very low ferritin concentrations. Comparison with serum iron and transferrin parameters in patients with iron deficiency demonstrated that serum ferritin concentrations might be subnormal in a majority of patients with otherwise normal iron indices. Up to 70% of the ferritin in serum of normal subjects could bind to concanavalin A-Sepharose, indicating its glycoprotein nature. It is concluded that our serum ferritin radioimmunoassay gave reliable results and was useful in the laboratory diagnosis of latent iron-deficiency and in the analysis of the heterogeneity of serum ferritin.

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The labelling of proteins to high specific radioactivities by conjugation to a 125I-containing acylating agent. Application to the radioimmunoassay

Cited by 3,217 publications

References 19 publications

Human complement subcomponent C2: purification and proteolytic cleavage in fluid phase by C1̄s, C1̄r₂–C1̄s₂ AND 1̄

Human complement subcomponent C2: purification and proteolytic cleavage in fluid phase by C1̄s, C1̄r₂–C1̄s₂ AND 1̄

Role of the C‐terminus in the activity, conformation, and stability of interleukin‐6

Measurement of ferritin in serum: Application in Diagnostic use

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The labelling of proteins to high specific radioactivities by conjugation to a 125I-containing acylating agent. Application to the radioimmunoassay

Cited by 3,217 publications

References 19 publications

Human complement subcomponent C2: purification and proteolytic cleavage in fluid phase by C1̄s, C1̄r2–C1̄s2 AND 1̄

Human complement subcomponent C2: purification and proteolytic cleavage in fluid phase by C1̄s, C1̄r2–C1̄s2 AND 1̄

Role of the C‐terminus in the activity, conformation, and stability of interleukin‐6

Measurement of ferritin in serum: Application in Diagnostic use

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Product

Resources

About

Human complement subcomponent C2: purification and proteolytic cleavage in fluid phase by C1̄s, C1̄r₂–C1̄s₂ AND 1̄

Human complement subcomponent C2: purification and proteolytic cleavage in fluid phase by C1̄s, C1̄r₂–C1̄s₂ AND 1̄