Section 1.

Introduction to
Molecular Diagnostics
 Goal

To introduce essential concepts in molecular

diagnostics that impact on the identification of
novel markers of human diseases and to
develop and apply useful molecular assays to
monitor disease, determine appropriate
treatment strategies, and predict disease
outcomes.
 Outline

• Definition of molecular diagnostics

• Impact of molecular diagnostics in human
disease
• Molecular pathology in laboratory medicine
• Molecular biology timeline
• Genomic technologies and applications
• Future prospects for molecular diagnostics
 Molecular Diagnostics

The use of molecular biology techniques to

expand scientific knowledge of the natural
history of diseases, identify people who are at
risk for acquiring specific diseases, and
diagnose infectious and other human diseases
at the nucleic acid level.
Guiding Principles for the Discovery
of Novel Molecular Markers
of Human Disease

Identification of novel molecular markers of

human disease will facilitate the development
of useful molecular assays for detection,
diagnosis, and prediction of disease outcomes.
 Molecular Diagnostics

• Monitor diseases more accurately

– Allows for early treatment and better patient
care
• Determine most appropriate treatment
– Reduces or eliminates unnecessary
treatment
– Reduces or eliminates inadequate treatment
– Yields greater cost effectiveness
• Reduce patient morbidity and mortality
Practical Applications of Molecular
Diagnostics in Laboratory Medicine

• Diagnostic
• Prognostic
• Therapeutic
• Predictive
Applications of Genomic Technologies
in Laboratory Medicine

• Anatomic pathology
• Chemistry/toxicology
• Genetics
• Hematopathology/oncology
• Infectious diseases
• Transfusion medicine/identity
 Molecular Pathology
A Universal Discipline of Laboratory Medicine
INFECTIOUS
DISEASE
HEMATOLOGY

Molecular
Pathology

SOLID
TUMORS
IDENTITY
TESTING

GENETIC
DISEASE
 Practical Applications of
Molecular Diagnostics in
Clinical Laboratory Medicine
Molecular Genetics
• Single gene disorders
• Polygenic disorders
• Chromosomal disorders

Molecular Oncology
• Diagnostic testing
• Disease prognosis
• Determination of predisposition
 Practical Applications of
Molecular Diagnostics in
Clinical Laboratory Medicine (cont.)

Hematopathology
• Diagnostic testing
• Determination of clonality

Identity Testing
• Parentage
• Clinical testing
 Practical Applications of
Molecular Diagnostics in
Clinical Laboratory Medicine (cont.)

Infectious Disease
• Qualitative and quantitative detection of
infectious agents
• Microbial identity testing
• Genotyping/drug resistance testing
Advances in the understanding of the structure
and chemistry of nucleic acids have facilitated
the development of technologies that can be
employed effectively in molecular
diagnostics.
The Molecular Biology Timeline

1865 Gregor Mendel, Law of Heredity

1866 Johann Miescher, Purification of DNA

1953 Watson and Crick, Structure of DNA

1970 Recombinant DNA Technology

1977 Sickle Cell Anemia Mutation

1980 In Vitro Amplification of DNA (PCR)

1995 The Human Genome Project

 Molecular Technologies in the
Clinical Laboratory

Blotting Techniques Electrophoretic Methods

Southern hybridization SSCP
Northern hybridization DGGE MDE

Amplification Techniques
PCR TMA
LCR NASBA
bDNA

DNA Sequencing
 Why are Nucleic Acids Important
in the Clinical Laboratory?

• Genetically-based diseases can be

diagnosed.
• Specificity can be controlled.
• Single base changes can be detected.
• Expression of gene product is not required.
• Targets can be amplified >105.
 Human Disease

Cause (etiology)

Mechanism (pathogenesis)

Structural alterations (morphologic/molecular)

Functional consequences (clinical significance)

 Understanding the Molecular
Pathogenesis of Human Disease
Enables Effective Utilization of
Molecular Assays
Diagnostic
• Distinguishing variants of human disease based
on presence of specific molecular markers
(chromosome translocations in lymphoma)
Prognostic
• Prediction of likely patient outcomes based on
presence of specific molecular markers (gene
mutations predicting poor clinical course in
cancer)
 Understanding the Molecular
Pathogenesis of Human Disease
Enables Effective Utilization of
Molecular Assays (cont.)
Therapeutic
• Prediction of response to specific therapies
based on presence of specific molecular
markers (gene mutations predicting poor drug
sensitivity in cancer)
 Future Prospects
for Molecular Diagnostics
Continued advances in the understanding of the
molecular basis of human disease will facilitate
development of new and useful molecular
diagnostics for a range of human disease
conditions.
Ultimately, identification of disease-causing genes
and/or genes that modify disease expression or
response to treatment will expand the number of
molecular diagnostics applications that will be
required for the basic clinical workup of patients.
 The Molecular Basis
of Human Disease

Genetic Lesions in Human Disease

• Identification of genetic markers
• Identification of disease-related genes
• Molecular targets for assay development
Characterization of Gene Sequences
• Facilitates characterization of disease-causing
mutations
• Molecular targets for assay development
 The Molecular Basis
of Human Disease (cont.)

Completion of the sequence of the human

genome will enable identification of all human
genes and establishment of disease-gene
relationships, facilitating development of
numerous new molecular assays.
 The Human Genome Project

• U.S. Government project coordinated by the

Department of Energy and the National
Institutes of Health
• Goals of the Human Genome Project
(1998–2003)
– to identify all of the genes in human DNA;
– to determine the sequences of the 3 billion bases
that make up human DNA;
– to create databases;
– to develop tools for data analysis; and
– to address the ethical, legal, and social issues that
arise from genome research.
 Beneficial Outcomes from
the Human Genome Project

• Improvements in medicine
• Microbial genome research
• DNA forensics/identity
• Improved agriculture and livestock
• Better understanding of evolution and human
migration
• More accurate risk assessment
 The Human Genome Project
Ethical, Legal, and Social Implications
• Use of genetic information
• Privacy/confidentiality
• Psychological impact and stigmatization
• Genetic testing
• Reproductive options/issues
• Education, standards, and quality control
• Commercialization
• Conceptual and philosophical implications
Molecular Diagnostics in Clinical Practice

The ultimate goal of the molecular diagnostics

laboratory is to provide molecular information
that will combine with and complement
information related to patient history and
symptomology, clinical laboratory results,
histopathological findings, and other diagnostic
information to provide a more sensitive, precise,
and accurate determination of disease diagnosis
and/or guidance toward appropriate and effective
treatment options.