Studies on chicken polyclonal anti-peptide antibodies specific for parathyroid hormone-related protein (1-36).
Chicken polyclonal antibodies were prepared against a synthetic peptide corresponding to the first 36 N-terminal amino acids of parathyroid hormone-related protein (PTHrP) by immunizing laying hens. Significant increases of antibodies to PTHrP were first detected after the second immunization. Production of anti-PTHrP egg yolk antibodies peaked 1-2 weeks after the second through sixth immunizations and declined over a period of 2-4 weeks. Polyclonal IgG (IgY) to PTHrP was purified from the egg yolks with high levels of PTHrP specific binding. The anti-PTHrP IgG was used to develop a radioimmunoassay for PTHrP that was able to detect 100 pg PTHrP ml-1 (23 pM) in conditioned cell culture medium. The anti-PTHrP IgG was bound to a solid phase and utilized to immunopurify iodinated [Tyr36]-PTHrP (1-36). Anti-PTHrP IgG inhibited the in vitro biologic activity of PTHrP as demonstrated by the inhibition of adenylate cyclase stimulation in a rat osteoblast-like cell line (ROS 17/2.8). The anti PTHrP IgG was immunopurified and utilized for immunohistochemical localization of PTHrP in canine skin. Chickens were advantageous in producing large amounts of high affinity, neutralizing antibodies to a highly conserved mammalian protein such as PTHrP. The antibodies will be useful to investigate the function and metabolism of PTHrP in vivo and in vitro.
Rosol
T J
TJ
Department of Veterinary Pathobiology, Ohio State University, Columbus 43210.
Steinmeyer
C L
CL
McCauley
L K
LK
Merryman
J I
JI
Werkmeister
J R
JR
Gröne
A
A
Weckmann
M T
MT
Swayne
D E
DE
Capen
C C
CC
D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D002645:Chickens / Q000276:immunology*; D002846:Chromatography, Affinity; D004530:Egg Yolk / Q000276:immunology; D007124:Immunoenzyme Techniques; D007074:Immunoglobulin G / Q000096:biosynthesis* / Q000302:isolation & purification; D044162:Parathyroid Hormone-Related Protein; D010446:Peptide Fragments / Q000276:immunology*; D011506:Proteins / Q000276:immunology*; D011863:Radioimmunoassay; D012867:Skin / Q000737:chemistry
D007074:Immunoglobulin G; D044162:Parathyroid Hormone-Related Protein; D010446:Peptide Fragments; D011506:Proteins; C066362:parathyroid hormone-related protein (1-36)amide, N(alpha)-(4-azido-2-nitrophenyl)-Ala(1)-Tyr(36)-