This article provides a comprehensive overview of microRNA (miRNA) profiling, including its methods, considerations, and applications. MiRNAs are small non-coding RNAs that regulate gene expression post-transcriptionally, and their profiling has gained popularity due to their potential as biomarkers in various diseases. The authors discuss the strengths and limitations of different profiling approaches, such as quantitative reverse transcription-PCR (qRT-PCR), hybridization-based methods, and high-throughput sequencing (RNA-seq). They also highlight the importance of sample processing and RNA extraction methods, which can significantly impact the results of miRNA profiling. The article further explores the applications of miRNA profiling in developmental biology, novel miRNA discovery, studying miRNA:mRNA and miRNA:protein interactions, and integrating miRNA profiles with other genomic datasets. Additionally, it discusses the use of miRNAs as biomarkers in diseases, including tissue-based biomarkers, circulating biomarkers, and forensic applications. The authors conclude by emphasizing the potential of miRNA profiling in developing clinically useful molecular diagnostic tests and the need for further advancements in sequencing technology to fully capitalize on the rich information content of miRNA profiles.This article provides a comprehensive overview of microRNA (miRNA) profiling, including its methods, considerations, and applications. MiRNAs are small non-coding RNAs that regulate gene expression post-transcriptionally, and their profiling has gained popularity due to their potential as biomarkers in various diseases. The authors discuss the strengths and limitations of different profiling approaches, such as quantitative reverse transcription-PCR (qRT-PCR), hybridization-based methods, and high-throughput sequencing (RNA-seq). They also highlight the importance of sample processing and RNA extraction methods, which can significantly impact the results of miRNA profiling. The article further explores the applications of miRNA profiling in developmental biology, novel miRNA discovery, studying miRNA:mRNA and miRNA:protein interactions, and integrating miRNA profiles with other genomic datasets. Additionally, it discusses the use of miRNAs as biomarkers in diseases, including tissue-based biomarkers, circulating biomarkers, and forensic applications. The authors conclude by emphasizing the potential of miRNA profiling in developing clinically useful molecular diagnostic tests and the need for further advancements in sequencing technology to fully capitalize on the rich information content of miRNA profiles.