Tissue microarray: an effective high-throughput method to study the placenta for clinical and research purposes.
OBJECTIVE
Tissue microarray (TMA) technology allows simultaneous examination of the expression of many molecular markers (protein, mRNA, DNA, etc.) with high-throughput. The application of this technology, to date, has been largely confined to the study of cancer. Placental pathology poses unique challenges because of the size of the organ, its complex anatomy, as well as its histological heterogeneity. The objective of this study was to assess the feasibility and efficiency of TMAs for immunohistochemistry and in situ hybridization of placental tissues.


METHODS
TMAs were constructed using an automated tissue arrayer. Standard 0.6-mm or 1-mm microarray needles were used. Villous parenchyma, basal plate, and chorioamniotic membranes were targeted in each block. Five mum-thick TMA sections underwent immunohistochemical analysis of both cytoplasmic and nuclear antigens using a panel of antibodies against a variety of cytoplasmic [cytokeratin-7, vascular endothelial growth factor (VEGF), and protein Z], membranous (endoglin), and nuclear (c-fos and c-jun) antigens. mRNA in situ hybridization for surfactant protein A (SP-A) and chromogenic in situ hybridization for the Y chromosome (DYZ1) were also performed.


RESULTS
Validation of TMA immunoreactivity demonstrated comparable results with corresponding whole sections. When a two-tiered scoring system (positive/negative) was employed, there was agreement between two and three cores and whole tissue sections (kappa>0.7). When a three-tiered scoring system (negative, weak-positive, or strong-positive) was used, the data from three cores showed the highest agreement with whole tissue sections (kappa >0.7). In situ hybridization experiments for mRNA and DNA were also successful in that the signals were readily detectable. Successful transfer from the donor block to the recipient block differed according to the anatomical compartment. The transfer efficiency of villous parenchyma, basal plate, and chorioamniotic membranes were 96.9% (875/903), 76.7% (115/150), and 75.4% (224/297), respectively.


CONCLUSIONS
TMA is a practical and effective tool for high-throughput molecular analysis of the human placenta. Duplicate and triplicate cores offer agreement with whole tissue sections for two-category distinction immunostaining. TMA also affords relevant results from in situ hybridization experiments for mRNA and DNA. The major advantages are the conservation of tissues and reagents, simultaneous comparison of molecular markers in different anatomical compartments of the placenta, and reduction of experimental error.
Richani
Karina
K
Perinatology Research Branch, National Institute of Child Health and Human Development, National Institutes of Health, Department of Health and Human Services, Bethesda, MD, USA.
Romero
Roberto
R
Kim
Yeon Mee
YM
Cushenberry
Enola
E
Soto
Eleazar
E
Han
Yu Mi
YM
Espinoza
Jimmy
J
Kim
Chong Jai
CJ
D015415:Biomarkers / Q000032:analysis; D035843:Biomedical Research / Q000379:methods; D004247:DNA / Q000032:analysis; D005240:Feasibility Studies; D005260:Female; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D017403:In Situ Hybridization / Q000379:methods; D046228:Microarray Analysis / Q000295:instrumentation / Q000379:methods*; D010920:Placenta / Q000166:cytology* / Q000473:pathology*; D011247:Pregnancy; D012333:RNA, Messenger / Q000032:analysis; D015203:Reproducibility of Results
D015415:Biomarkers; D012333:RNA, Messenger; D004247:DNA