Peptide Nucleic Acid (PNA) are DNA/RNA synthetic analogs with 2-((2-Aminoethyl) amino) acetic acid backbone. PNA has replaced DNA as a probe in various pre-existing techniques and currently is a prominent biomolecular tool for genetic diagnosis, cytogenetics, and pharmaceutical applications. PNAs physicobiochemical properties make them resistant to enzymatic cleavage and do not degrade in biological conditions. PNA partakes unique antisense and antigene properties, just due to its inhibitory effect on transcription and translation, and undergo complementary binding to RNA/DNA with high affinity and specificity. Hence, to date, many methods utilizing PNA for the detection and treatment of various diseases such as cancer, AIDS, etc. have been designed and developed. PNA is used for PCR reaction modulation/mutation, in fluorescent in-situ hybridization (FISH) and microarray as a probe, also in many in-vitro and in-vivo assays, and in the development of micro and nano-sized biosensor/chip/array technologies. Earlier reviews focused only on PNA properties, structure and modifications related to diagnostics and therapeutics; our review emphasizes not only on PNA properties but also on its potential applications in diagnosis and therapeutics, along with its prospects in biomedical applications.