High-Performance Liquid Chromatography of 125I-Labeled Mouse Epidermal Growth Factor Radioiodinated by Six Different Methods

1992 ◽  
Vol 38 (5) ◽  
pp. 681-686 ◽  
Author(s):  
C B Kienhuis ◽  
J J Heuvel ◽  
H A Ross ◽  
J A Foekens ◽  
T J Benraad
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
High Performance ◽  
Binding Capacity ◽  
Reversed Phase ◽  
Iodine Monochloride ◽  
Reversed Phase Hplc ◽  
Hplc Column ◽  
Epidermal Growth ◽  
Chloramine T

Abstract Six different procedures for radioiodination of mouse epidermal growth factor (EGF) all resulted in a heterogeneous 125I-labeled EGF preparation, as analyzed by reversed-phase HPLC. EGF preparations that had been iodinated with Chloramine T, lodogen, or lodo-beads were found mainly to consist of oxidized 125I-labeled EGF moieties. In contrast, the heterogeneous 125I-labeled EGF preparations obtained by using iodine monochloride, Protag-125, or lactoperoxidase-glucose oxidase-coupled beads (Enzymobeads) contained insignificant amounts of oxidized EGF entities. Ligand equivalence analysis (LEA) of distinct HPLC column fractions, obtained after preparative separation of Chloramine T-125I-labeled EGF, showed that the receptor-binding affinity of the tracer in all subfractions was less than the affinity of unlabeled EGF. This implies that HPLC purification of these 125I-labeled EGF preparations does not yield 125I-labeled EGF preparations with ligand equivalence. However, all but one HPLC column fraction of Enzymobeads-125I-labeled EGF showed ligand equivalence. Despite the small amount of the nonequivalent component in the Enzymobeads-labeled tracer, the nonchromatographed 125I-labeled EGF preparation showed ligand equivalence. No significant differences were observed in the maximal binding capacity of the different 125I-labeled EGF preparations.

scholarly journals The Fate of Tritiated rm-Epidermal Growth Factor in the Sheep: Validation of the Labelling Procedure and Rate of Tissue Clearance

1988 ◽  
Vol 41 (4) ◽  
pp. 539 ◽  
Author(s):  
IH O'Keefe ◽  
LF Sharry ◽  
BA Panaretto
Keyword(s):  
Amino Acid ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
High Performance ◽  
Fast Atom Bombardment ◽  
Parent Compound ◽  
Reversed Phase ◽  
Labelling Procedure ◽  
Epidermal Growth ◽  
Hydrogen Substitution

Plasmid-derived recombinant mouse epidermal growth factor, rm-EGF, was purified by ion pair reversed phase high performance liquid chromatography. The product peak (termed rm-a-EGF) was characterized by physicochemical techniques including fast atom bombardment mass spectrometry, high field proton magnetic resonance and amino acid sequencing (amino acid arrangement and composition). The rm-a-EGF was tritiated, labile tritium removed by lyophilization: and the product purified and characterized as for the parent compound to yield a compound identical to rm-a-EGF except for the isotopic hydrogen substitution. Label stability was validated by lyophilization of samples, especially urine.

Epidermal growth factor in gingival crevicular fluid and its binding capacity in inflamed and non-inflamed human gingiva

1996 ◽  
Vol 41 (7) ◽  
pp. 719-724 ◽  
Author(s):  
Kuang-Min Chang ◽  
Nina Lehrhaupt ◽  
Louis M. Lin ◽  
Jian Feng ◽  
Chi-Ying Wu-Wang ◽  
...  
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Gingival Crevicular Fluid ◽  
Binding Capacity ◽  
Human Gingiva ◽  
Epidermal Growth ◽  
Crevicular Fluid

Human serum and plasma have different sources of epidermal growth factor

1990 ◽  
Vol 259 (3) ◽  
pp. R545-R548 ◽  
Author(s):  
A. Lev-Ran ◽  
D. L. Hwang ◽  
D. S. Snyder
Keyword(s):  
Bone Marrow ◽  
Platelet Count ◽  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Molecular Mass ◽  
High Performance ◽  
Regression Line ◽  
Bone Marrow Ablation ◽  
Epidermal Growth ◽  
Marrow Ablation

Epidermal growth factor (EGF) was determined by radioimmunoassay in serum, plasma, and urine of 23 patients undergoing ablative therapy followed by bone marrow transplantation. The difference between the serum and plasma values reflected the amount of EGF released from the platelets at the time of blood coagulation. Platelet-derived EGF strongly correlated with platelet count (r + 0.850, P less than 0.0001), and the intercept of the regression line was very close to zero; one platelet contained approximately 2.5 x 10(-18) g EGF. Correspondingly, when the platelet count dropped after bone marrow ablation from 222 +/- 97 to 33 +/- 13 x 10(9)/l, the serum EGF decreased from 603 +/- 222 to 65 +/- 41 pg/ml (P less than 0.0001). Plasma EGF content did not correlate with the platelet count and did not change significantly after bone marrow ablation (before and after the ablation, correspondingly, 290 +/- 80 and 332 +/- 99 pg/ml, P = 0.194). High-performance liquid chromatographic fractionation of serum and plasma showed different molecular mass distribution of EGF-immunoreactive fractions. The main molecular mass components of the plasma EGF did not change after bone marrow ablation. Urine excretion remained unchanged (320 +/- 133 and 314 +/- 173 pmol EGF/mmol creatinine). We conclude that whereas platelets are the source of serum EGF, the origin of plasma EGF is different and the search of its origin is warranted.

scholarly journals 125I-labeled human epidermal growth factor. Binding, internalization, and degradation in human fibroblasts.

1976 ◽  
Vol 71 (1) ◽  
pp. 159-171 ◽  
Author(s):  
G Carpenter ◽  
S Cohen
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Energy Production ◽  
Binding Capacity ◽  
Human Fibroblasts ◽  
Metabolic Energy ◽  
Acid Extraction ◽  
Native Form ◽  
Human Epidermal Growth Factor ◽  
Epidermal Growth

125I-labeled human epidermal growth factor (hEGF) binds in a specific and saturable manner to human fibroblasts. At 37 degrees C, the cell-bound 125I-hEGF initially may be recovered in a native form by acid extraction; upon subsequent incubation, the cell-bound 125I-hEGF is degraded very rapidly, with the appearance in the medium of 125I-monoiodotyrosine. At 0 degrees C, cell-bound 125I-hEGF is not degraded but slowly dissociates from the cell. The data are consistent with a mechanism in which 125I-hEGF initially is bound to the cell surface and subsequently is internlized before degradation. The degradation is blocked by inhibitors of metabolic energy production (azide, cyanide, dinitrophenol), some protease inhibitors (Tos-Lys-CH2Cl, benzyl guanidobenzoate), a lysosomotropic agent (chloroquine) various local anesthetics (cocaine, lidocaine, procaine), and ammonium chloride. After the binding and degradation of 125I-hEGF the fibroblasts are no longer able to rebind fresh hormone. The binding capacity of these cells is restored by incubation in a serum-containing medium; this restoration is inhibited by cycloheximide or actinomycin D.

Sign in / Sign up

Export Citation Format

Share Document