Cellular Pathology in recent years has
become more closely involved in the direct management of patients with the
introduction of molecular technologies and targeted therapies. Through this, we
have seen the introduction of specialist pathology. These concepts and the key
technologies that are influencing clinical practice today have been introduced.
It showed that how clinical practice has been affected by these respective
technologies and how further development will influence the practice and
delivery of cellular pathology, which will impact on the patient through
targeted therapeutics and diagnostics.
There are many types of advanced techniques
in cellular pathology.
3,000,000 base pairs of nucleotide that
is 0.1% of the genomes are unique in all human being. This speciality in the base
sequences occur in repetitive DNA also known as satellite DNA as well as in
genes. Various small peaks are formed on the DNA which gives rise to
polymorphism due to density gradient configuration in the satellite DNA. Variable number tandem repeats (VNTR)
is one of the main satellite DNA having high degree of polymorphism. Since a
child receive 50% of the DNA from its father and the other 50% from his mother,
so the number VNTRs at a particular area of the DNA of the child will be
different may be due to insertion, deletion or mutation in the base pairs. As a
result, every individual has a distinct composition of VNTRs and this is the
main principle of DNA fingerprinting.
profiling, as already indicated, has application in a broad cross section of
disciplines, including human forensic science, diagnostic medicine, family
relationship analysis, animal and plant sciences, and wildlife forensic
science. DNA profiling is applicable in a number of areas in medicine including twin zygosity testing , bone marrow
transplantation marker analysis, detection of DNA changes in tumors, indication
of possible contamination of fetal by maternal tissue in chorionic villus
analysis, pathogen identification and paternity
testing where family studies are being performed for antenatal diagnosis of
The principle of PCR involves the primer mediated enzymatic amplification
of DNA. The ability of DNA polymerase to synthesize new strand of DNA
complementary to the offered template strand are by using PCR. DNA polymerase
can add a nucleotide only onto a preexisting 3?-OH group to add the first
nucleotide, so the primer is needed. DNA polymerase then elongated its 3 end by
adding more nucleotides to generate an extended region of double stranded DNA.
There are many applications of PCR
including to test the presence of genetic disease mutation such as cystic
fibrosis, hemoglobinopathies or other inborn errors of metabolism. It can used
to study the alteration to oncogene that causes cancer. It is also a tool used
in genetic fingerprinting. In a crime investigation, there may only be tiny DNA
sample to work with to identify anyone from the million.
Real-time PCR (RT-PCR):
Real-time PCR is the continuous
collection of fluorescent signal from one or more polymerase chain reactions
over a range of cycles. PCR
products is enabled by the inclusion of a fluorescent reporter molecule in each
reaction well that yields increased fluorescence with an increasing amount of
product DNA by real-time detection. The fluorescence
employed for this purpose include DNA-binding dyes and fluorescently labeled
sequence-specific primers or probes. The fluorescence signal as amplification
occurs are monitor by using specialized thermal cyclers equipped with
fluorescence detection modules. The measured fluorescence is proportional to
the total amount of amplicon; the change in fluorescence over time is used to
calculate the amount of amplicon produced in each cycle. RT-PCR
assays have become the best of choice for the rapid and sensitive determination
and quantitation of nucleic acid in various biological samples, with diverse
applications such as gene expression analysis, the detection of genetically
modified organisms in food, and cancer phenotyping.
tool of choice for the rapid and sensitive determination and quantitation of
nucleic acid in various biological samples is real-time PCR assays. The detection
of genetically modified organisms in food, gene expression analysis and cancer
phenotyping are main applications of real-time PCR. In research laboratories,
RT-PCR assays are widely used for the quantitative measurement of gene copy
number (gene dosage) in transformed cell lines or the presence of mutant genes.
It can be used to precisely quantitate changes in gene expression, for example,
an increase or decrease in expression in response to different environmental
conditions or drug treatment, by measuring changes in cellular mRNA levels.