ABSTRACT
BACKGROUND: Karyotype determination has a central role in the genetic workup of pregnancy loss, as aneuploidy (trisomy and monosomy) and polyploidy (triploidy and tetraploidy) are the cause in at least 50% of first trimester, 25% of second trimester, and 11% of third trimester miscarriages. There are several limitations with the current approaches of obtaining a karyotype using traditional cytogenetics, fluorescence in situ hybridization with a limited number of probes, and chromosomal microarray. These include culture failure, incomplete results, lower sensitivity, and longer reporting time. METHODS: To overcome current limitations, a novel molecular assay is developed with a Standard Resolution Interphase Chromosome Profiling probe set which is a variation of the recently developed High Resolution probe set. It generates a molecular karyotype that can detect all major changes commonly associated with pregnancy loss. Initial familiarization of signal patterns from the probe set was used, followed by validation of the method using 83 samples from miscarriages in a blind study from three different laboratories. Finally, the clinical utility of the method was tested on 291 clinical samples in two commercial reference laboratory settings on two different continents. RESULTS: The new molecular approach not only identified all the chromosome changes observed by current methods, but also significantly improved abnormality detection by characterizing derivative chromosomes and finding subtle subtelomeric rearrangements, balanced and unbalanced. All Robertsonian translocations were also detected. The abnormality rate was 54% on clinical samples from commercial laboratory 1 and 63% from laboratory 2. CONCLUSION: The attributes of this method make it an ideal choice for the genetic workup of miscarriages, namely (1) near 100% successful results, (2) greater sensitivity than conventional chromosome analysis or FISH panels, (3) rapid reporting time, and (4) favorable comparisons with chromosomal microarray.
Subject(s)
Cytogenetic Analysis/methods , Cytogenetics/methods , Abortion, Spontaneous/genetics , Chromosome Aberrations , Female , Humans , In Situ Hybridization, Fluorescence/methods , Interphase/genetics , Karyotype , Karyotyping/methods , Monosomy/diagnosis , Pregnancy , Prenatal Diagnosis/methods , Sensitivity and Specificity , Tetrasomy/diagnosis , Trisomy/diagnosisABSTRACT
CONTEXT: - Chromosome analysis on bone marrow or peripheral blood samples fails in a small proportion of attempts. A method that is more reliable, with similar or better resolution, would be a welcome addition to the armamentarium of the cytogenetics laboratory. OBJECTIVE: - To develop a method similar to banded metaphase chromosome analysis that relies only on interphase nuclei. DESIGN: - To label multiple targets in an equidistant fashion along the entire length of each chromosome, including landmark subtelomere and centromere regions. Each label so generated by using cloned bacterial artificial chromosome probes is molecularly distinct with unique spectral characteristics, so the number and position of the labels can be tracked to identify chromosome abnormalities. RESULTS: - Interphase chromosome profiling (ICP) demonstrated results similar to conventional chromosome analysis and fluorescence in situ hybridization in 55 previously studied cases and obtained useful ICP chromosome analysis results on another 29 cases in which conventional methods failed. CONCLUSIONS: - ICP is a new and powerful method to karyotype peripheral blood and bone marrow aspirate preparations without reliance on metaphase chromosome preparations. It will be of particular value for cases with a failed conventional analysis or when a fast turnaround time is required.
