Hemophilia A (HA) due to defective F8 is an X-linked hereditary disorder characterized by bleeding of variable severity through mild, moderate to severe owing to large range of mutations [1]. All kind of mutations, except repeats have been reported for HA, in total up to 2370 [2]. A preliminary study was conducted in our lab for identification of mutations in F8 gene in HA patients with an emphasis on study of molecular pathology of Pakistani HA patients and their correlation with clinical manifestations. For this study, all exons of F8 gene were selected. Samples were collected from 62 HA patients from all over the Pakistan and their clinical history was recorded (only patients frequently visiting medical centers for frequent treatment of F8 were selected for the study). Their genomic DNA was separated by standard procedures. Specific primers (Figure-1) were designed using "Primer3" (http://biotools.umassmed.edu/bioapps/primer 3_www.cgi) to amplify the selected portions of the gene; amplification products were sequenced by ABI 310 and ABI 3100 sequencer (Applied Biosystems). The sequencing results were visualized using "Chromas2.33" software (Applied Biosystem) and mutations were detected using "BLAST" software available on the NCBI website (http:balst.ncbi.nlm.nih.gov/Blast.cgi? PROGRAM=blastnB 2 point mutations) were detected in four sporadic patients, all from different ethnic backgrounds. Single T deletion at Exon 7 within the A1 domain clearly represents a frame-shift change disrupting protein structure and function resulting in severe manifestation of the disease. A missense point mutation in the A3 domain occurs at codon 1907 at nucleotide number 5720 replacing Serine with Isoleucine and confers a moderate type of severity, although both amino-acids have different properties: serine being polar and acidic and Isoleucine a nonpolar and basic compound. A nonsense point-mutation was available in two unrelated patients in the C3 domain in exon 26 manifesting moderate hemophilic conditions. Beside these mutations, 27 common SNPs were also detected among exons of F8 for all the patients (Table-2). Their allelic data and accession numbers were collected from Ensembl Genome Browser [3]. The results will not only form the basis of an enlarged study but also will be useful in the genetic and molecular analysis/diagnosis and genetic counseling of classical hemophilia in Pakistan.