Peptide Nucleic Acid (PNA) are DNA/RNA synthetic analogs with 2-([2-aminoethyl] amino) acetic acid backbone. They partake unique antisense and antigene properties, just due to its inhibitory effect on transcription and translation; they also undergo complementary binding to RNA/DNA with high affinity and specificity. Hence, to date, many methods utilizing PNA for diagnosis and treatment of various diseases namely cancer, AIDS, human papillomavirus, and so on, have been designed and developed. They are being used widely in polymerase chain reaction modulation/mutation, fluorescent in-situ hybridization, and in microarray as a probe; they are also utilized in many in-vitro and in-vivo assays and for developing 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 on PNA properties and synthesis along with its potential applications in diagnosis and therapeutics. Furthermore, prospects in biomedical applications of PNAs are being discussed in depth.