MICROSCOPY AND MOLECULAR TECHNIQUES
Learning outcomes of the course unit
The course aims to provide the methodology and the laboratory techniques employed by pathologists in the clinical practice and the research.
The main microscopic and molecular methods used in modern pathology will be discussed along with their diagnostic applications.
The acquired knowledge will allow the student to understand the variety of methodological approaches that can be applied to the study of pathological specimens. It will also allow the student to choose the more appropriate technique to the resolution of an experimental or diagnostic problem.
The student will be asked to integrate the notions provided by the course with previous knowledge in anatomy, molecular biology physiology and general pathology to the understanding of:
• the specificity and the relevance of a tissue approach to the study of disease;
• generalities on the molecular basis of cancer;:
• the physical, chemical and molecular premises to the use of the different morphological and molecular techniques;
• their main operative procedures;
• the logistic, instrumental and quality control requirements for their use in the diagnostic laboratory;
• their indications of use and main diagnostic applications.
At the end of the term, the student will be able to interact with other professional figures (clinicians, biotechnologists, pathologists, etc.) in the:
• choice of the more appropriate technique to solve a given experimental or diagnostic problem;
• critical interpretation of the results;
• integration of a tissue analysis approach in wider strategies of diagnosis and cure.
Basic notions of histology, physiology, molecular biology and general pathology.
Course contents summary
The first part of the course will introduce the morphological and molecular approaches to the study of pathological cells and tissues. Basic notions on hallmarks and molecular basis of cancer will also be provided. Finally, target molecular therapy of cancer will be discussed.
The second part of the course will examine the basic procedures of histopathology and the morphological techniques including histochemical stains, immunohistochemistry, in situ hydridization, electron microscopy, advacnced microscopy techniques, automation in in Pathology, digital slides and their applications.
The third part of the course will be dedicated to the main molecular techniques that can be applied to the research and diagnosis of human tissues including: PCR and its variants, DNA sequencing, techniques of point mutation analysis, LOH, clonality assays, analysis of DNA methylation, expression arrays, CGH array and tissue microarrays.
• Surgical pathology: from diagnosis to research.
• Introduction to the methodology and technical approaches to the morphological study of pathologic tissues.
• Hallmarks of cancer.
• Laboratory and techniques for light and electron microscopy: grossing and sampling, fixation, inclusion, routine stains, special histochemical stains.
• Immuno-histochemical techniques, immuno-fluorescence and ‘in situ’ hybridization, applications to clinical practice and research.
• Advanced microscopy techniques and introduction to digital pathology
• Automation in Pathology
• Extraction and purification of bio molecules.
• Analysis of bio molecules: quantification, study of integrity and quality.
• Amplification of nucleic acids: PCR, quantitative PCR, reverse transcriptase PCR, whole genome amplification.
• Epigenetic analysis in the study of tumours: test for the analysis of methylation status in the clinical practice.
• Approach to familiar colorectal cancer: hereditary syndromes(FAP, HNPCC),the Bethesda panel.
• Oncogenes, tumor suppressor genes and the "mismatch repair" system in colorectal-cancer.
• Analysis of microsatellites (LOH & MIN), immunostains and, sequencing.
• Techniques for the detection of gene mutations.
• Target therapy
• Molecular biology techniques for the detection of drug targets.
• DNA and RNA arrays in the study of neoplasia
• First and second generation DNA sequencing
• Analysis of clonality
• Molecular approaches to the analysis of the minimal residual disease.
Selected references and teaching material provided by teachers.
Lessons, seminars and laboratories will be given during the course. Texts, diagrams, clinical examples and a rich apparatus of gross and microscopic images will be used to guide the student into the understanding of the characteristics and application of the different techniques.
Study groups will be formed in which the students will be called to study, understand and critically review a relevant topic of the more recent scientific literature. This work will be translated into an oral presentation.
Assessment methods and criteria
An oral interview will be used to assess the reaching of learning targets. Questions will be aimed to assess knowledge and understanding of basic notions. Questions will be asked concerning the use the various techniques in different experimental and diagnostic scenarios to assess the ability of the student to translate knowledge into appropriate actions and behaviors in relevant clinical situations.
The final results of the examination will consider both the oral interview and the group presentation.
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