Related Experiment Video
Updated: Aug 3, 2026
Cerebrospinal Fluid MicroRNA Profiling Using Quantitative Real Time PCR
Published on: January 22, 2014
Identification and validation of reference genes of circulating microRNAs for use as control in gestational
Ingrid de Siqueira Pereira1, Allecineia Bispo da Cruz1, Marta Marques Maia1
1Centro de Parasitologia e Micologia, Instituto Adolfo Lutz, Sao Paulo, Brazil; Programa de Pós-Graduação em Ciências da Coordenadoria de Controle de Doenças da Secretaria de Estado da Saúde de São Paulo, Brazil.
Abstract:
Toxoplasmosis causes serious harm to the fetus, as tachyzoite dissemination, during pregnancy in women developing the primo-infection. The microRNAs (miRNAs) are small non-coding RNAs, which have regulatory roles in cells by silencing messenger RNA. Circulating miRNA are promising biomarkers for diagnosis and prognosis of numerous diseases. The miRNAs levels are estimated by quantitative real-time PCR (qPCR), however, the relative quantification of each miRNA expression requires proper normalization methods using endogenous miRNAs as control. This study analyzed the expression of three endogenous miRNAs (miR-484, miR -423-3p and miR-26b-5p) for use as normalizers in future studies of target miRNAs for gestational toxoplasmosis (GT). A total of 32 plasma samples were used in all assays divided in 21 from women with GT and 11 from healthy women. The stability of each endogenous miRNA was evaluated by the algorithm methods RefFinder that included GeNorm, Normfinder, BestKeeper and comparative delta-CT programs. The miR-484 was the most stably gene, and equivalently expressed in GT and NC groups. These results contribute to future studies of target miRNAs in clinical samples of women with gestational toxoplasmosis.
Insights
This study identified miR-484 as a stable endogenous reference gene for quantifying microRNAs in gestational toxoplasmosis (GT) plasma samples. This finding supports accurate biomarker development for diagnosing and monitoring fetal toxoplasmosis.
Area of Science:
- Biochemistry
- Molecular Biology
- Genetics
Background:
- Gestational toxoplasmosis (GT) poses significant fetal risks due to parasite dissemination.
- Circulating microRNAs (miRNAs) show promise as diagnostic and prognostic biomarkers.
- Accurate miRNA quantification requires reliable endogenous reference genes for normalization in quantitative real-time PCR (qPCR).
Purpose of the Study:
- To evaluate the stability of three endogenous miRNAs (miR-484, miR-423-3p, miR-26b-5p) as normalizers for miRNA expression studies in gestational toxoplasmosis.
- To identify the most stable miRNA for accurate quantification in clinical samples.
Main Methods:
- Analysis of plasma samples from 21 women with GT and 11 healthy controls.
- Evaluation of miRNA stability using RefFinder algorithm, incorporating GeNorm, Normfinder, BestKeeper, and comparative delta-CT methods.
- Quantitative real-time PCR (qPCR) for miRNA expression analysis.
Main Results:
- miR-484 demonstrated the highest stability among the tested endogenous miRNAs.
- miR-484 exhibited equivalent expression levels in both gestational toxoplasmosis and normal control groups.
- The stability of miR-484 was confirmed across multiple analytical algorithms.
Conclusions:
- miR-484 is a suitable and stable endogenous reference gene for normalizing miRNA expression in plasma samples from women with gestational toxoplasmosis.
- This finding facilitates future research on miRNA biomarkers for GT diagnosis and prognosis.
- Validated normalization strategies are crucial for reliable clinical applications of circulating miRNAs.
More Related Videos
10:37Perturbations of Circulating miRNAs in Irritable Bowel Syndrome Detected Using a Multiplexed High-throughput Gene Expression Platform
Published on: November 30, 2016
08:12Detection of a Circulating MicroRNA Custom Panel in Patients with Metastatic Colorectal Cancer
Published on: March 14, 2019
Related Concept Videos
01:22MicroRNAs
01:22MicroRNAs