A high-throughput method for quantifying gene expression data from early Drosophila embryos.
We describe an automated high-throughput method to measure protein levels in single nuclei in blastoderm embryos of Drosophila melanogaster by means of immunofluorescence. The method consists of a chain of specific algorithms assembled into an image processing pipeline. This pipeline transforms a confocal scan of an embryo stained with fluorescently tagged antibodies into a text file. This text file contains a numerical identifier for each nucleus, the coordinates of its centroid, and the average concentrations of three proteins in that nucleus. The central algorithmic component of the method is the automatic identification of nuclei by edge detection with the use of watersheds as an error-correction step. This method provides high-throughput quantification at cellular resolution.
Janssens
Hilde
H
Department of Applied Mathematics and Statistics, and Center for Developmental Genetics, Stony Brook University, Stony Brook, NY 11794-3600, USA.
Kosman
Dave
D
Vanario-Alonso
Carlos E
CE
Jaeger
Johannes
J
Samsonova
Maria
M
Reinitz
John
J
D000465:Algorithms; D000818:Animals; D018951:Antigens, CD20 / Q000378:metabolism; D004330:Drosophila / Q000196:embryology* / Q000235:genetics*; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism; D004625:Embryo, Nonmammalian / Q000378:metabolism; D019085:Fluorescent Antibody Technique, Direct; D018507:Gene Expression Regulation, Developmental; D017344:Genes, Insect; D006454:Hemoglobins / Q000378:metabolism; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D007091:Image Processing, Computer-Assisted; D019476:Insect Proteins / Q000235:genetics / Q000378:metabolism; D018613:Microscopy, Confocal; D013997:Time Factors; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
D018951:Antigens, CD20; D029721:Drosophila Proteins; D006454:Hemoglobins; D018398:Homeodomain Proteins; D019476:Insect Proteins; D014157:Transcription Factors; C052837:eve protein, Drosophila