A robust immunoassay for pregnancy-associated plasma protein-A2 based on analysis of circulating antigen: establishment of normal ranges in pregnancy

Abstract

Pregnancy-associated plasma protein-A (PAPP-A) and PAPP-A2, two homologous metzincin metalloproteases, are both tightly linked to regulation within the insulin-like growth factor (IGF) system because of their specific cleavage of IGF binding proteins. Recent studies suggest that PAPP-A may be involved in clinical conditions related to unwanted cellular growth, and the circulating levels of PAPP-A is an established biomarker in prenatal screening for chromosomal abnormalities. Microarray data indicate that PAPP-A2 has potential as a biomarker for pre-eclampsia. However, well-characterized immunological methods of quantification are not available. We therefore developed monoclonal antibodies against recombinant PAPP-A2. The antibodies were epitope mapped against recombinantly expressed chimeras between PAPP-A2 and PAPP-A. Furthermore, circulating PAPP-A2 was immunoaffinity purified and characterized by sequence analysis and mass spectrometry. Unlike PAPP-A, PAPP-A2 is a noncovalent dimer in which each subunit of 1558 amino acids originates from all of the 22 predicted coding exons. A previously hypothesized variant (PAPP-E) does not exist, but low amounts of a C-terminally truncated PAPP-A2 variant was detected. A sensitive and robust ELISA for full-length PAPP-A2 was developed and used to establish normal ranges of PAPP-A2 through pregnancy. The functional sensitivity of this ELISA at 20% CV was 0.08 ng/ml, and the serum concentration of PAPP-A2 was found to increase during pregnancy in agreement with placental synthesis. The existence of this assay will enable an assessment of the biomarker potential of PAPP-A2 in pre-eclampsia as well as other clinical conditions.

Pregnancy-Associated Plasma Protein A2 in Hemodialysis Patients: Significance for Prognosis

“…production by placenta and increase during pregnancy, there are also some differences. First, PAPP-A increase is much more pronounced compared to the increase of PAPP-A2 [9, 19, 23]. Second, in case of Down syndrome or pregnancy pathologies there is early decrease of PAPP-A [7, 30] while PAPP-A2 increases [25-29].…”

“…46% of its amino acid residues with PAPP-A and it is synthesized as a prepro-protein [1]. Unlike PAPP-A, PAPP-A2 circulates as a noncovalent dimer [18, 19] and each subunit consists of 1558 amino acids (220kDa) [19]. Additionally, a C-terminally truncated PAPP-A2 variant was detected (less than 5% of PAPP-A2) [19].…”

“…Unlike PAPP-A, PAPP-A2 circulates as a noncovalent dimer [18, 19] and each subunit consists of 1558 amino acids (220kDa) [19]. Additionally, a C-terminally truncated PAPP-A2 variant was detected (less than 5% of PAPP-A2) [19]. PAPP-A2 cleaves IGFBP-5 and this cleavage is IGF-independent [1].…”

“…The average level at the end of pregnancy is 500-fold higher than the levels outside pregnancy [19]. This difference is much less pronounced compared to that of PAPP-A, which is more than 10,000 fold [9, 23].…”

Surface plasmon resonance biosensor for detection of pregnancy associated plasma protein A2 in clinical samples

“…PAPP-A2 plays multiple roles in fetal development and post-natal growth and it is believed to be connected to disturbances during pregnancy (e.g., pre-eclampsia or HELLP syndrome [1]). Although the physiological functions of PAPP-A2 are still under investigation, it is believed that they could be related to PAPP-A and, furthermore, that functions of PAPP-A2 and PAPP-A are linked to one another.…”