Download - Advanced diagnostic tecniques

04/08/2023 1

Advanced DiagnosticTechnologies in Public

Health

Dr Abhay Dhanorkar

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Scope

• Introduction• IPHS standards for availability of laboratory services at various hospitals.• Classification of diagnostics tests• Immunopathology• Molecular Biology• Clinical Chemistry• Haematology• Cytopathology• Histopathology• Radiology• Electrophysiology• Endoscopy• Summary

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Introduction

• A diagnostic test is any kind of medical test performed to aid in the diagnosis or detection of disease.

• It may be used to diagnose diseases or measure the progress or recovery from disease or confirm that a person is free from disease.

• Some medical tests are parts of a simple physical examination which require only simple tools in the hands of a skilled practitioner and can be performed in an office environment.

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• Some other tests require elaborate equipment used by medical technologists or the use of a sterile operating theatre environment.

• Some tests require samples of tissue or body fluids to be sent off to a pathology lab for further analysis.

• Some simple chemical tests such as urine pH, can be measured directly in the physicians office.

Introduction contd……

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• However, with advancements in health care, a large array of diagnostic tests have become available. Various modern (and often, expensive) diagnostic modalities are often routinely used in medical world.

• According to IPHS standards stated as a part of NRHM, the some laboratory facilities should be available at various levels.

Introduction contd……

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IPHS standards for availability of laboratory services at 31 to 50 bedded hospitals

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For 51-100 Bedded (All as in 31-50 bedded with the following additional

facilities)

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facilities)

Speciality Diagnostic Services / Tests

Endoscopy Colonoscopy, Bronchoscopy, Arthroscopy, Laparoscopy (Diagnostic), Colposcopy, Hysteroscopy

Serology RA factor test CPK, Lepto spirosis (Rapid test / ELISA)

Radiology HSG, Ultrasonography Colour Doppler, Spiral CT scan, MRI 0.5 TESSLA

Biochemistry Glycosylated Hemoglobin, Blood gas analysis

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facilities)

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For >300 Bedded (All as in 201-300 bedded with the following additional

facilities)Speciality Diagnostic Services

Radiology Angiography, Multislice CT, Multislice MRI

Biopsy Bone Marrow Biopsy,Pleural Biopsy, Pericardial tapping

Endoscopic Specialised Procedures and Diagnostic

Gastroscopy (Oesophagus, stomach, deudenum), Sigmoid, Laproscopy and

Colonoscopy, Bronchoscopy and Foreign Body Removal

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Classification

• The various modern diagnostic technologies can be classified as– Immunopathology– Molecular Biology– Clinical Chemistry– Haematology– Cytopathology– Histopathology– Radiology– Electrophysiology– Endoscopy

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Diagnostic Technologies in Immunopathology

• Antibodies are produced in response to antigenic stimulation.

• Immunoassays (antigen antibody reactions) can be used for detection of either antigens or antibodies. For detecting antigens, the corresponding specific antibody should be used as one of the reagents and vice versa.

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The technologies available are –• Precipitation immunoassays : It is based on the occurrence of

precipitation when large complexes of antigens and antibodies combine to form an insoluble lattice.Ex. :- Widal test and the Weil-Felix test.

• Particle immunoassay : Here specific antigens are coated onto a particle and on reaction with antibody under test, the agglutination is made more visible. Reverse agglutination where the antibody is coated onto the particle is also performed. The particle used may be RBC, latex or gelatin. Ex. :- 1) TPHA (Treponema pallidum Hemagglutination).

2) Reverse passive hemagglutination (RPHA) for hepatitis B surface antigen (HBsAg). 3) Latex agglutination is used in measurement of hCG for qualitative pregnancy tests

Immunopathology contd…

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Radioimmunoassay : • Radioimmunoassay (RIA) is used to test antigens

(e.g.hormone levels in the blood). • It is extremely sensitive and extremely specific, but requires a

sophisticated apparatus and is costly. It also requires special precautions, since radioactive substances are used.

• Therefore, today it has been largely supplanted by the ELISA method, where the antigen-antibody reaction is measured using colorometric signals instead of a radioactive signal.

• To perform a radioimmunoassay, a known quantity of an antigen is made radioactive, frequently by labeling it with gamma-radioactive isotopes of iodine attached to thyrosine.


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• This radiolabeled antigen is then mixed with a known amount of antibody for that antigen and as a result, the two chemically bind to one another.

• Then, a sample of serum from a patient containing an unknown quantity of that same antigen is added. This causes the unlabeled (or “cold”) antigen from the serum to compete with the radiolabeled antigen for antibody binding sites.

Applications in public health : Ex. Thyroid hormone levels in goiter endemic areas.


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Enzyme Linked Immunosorbent Assay (ELISA) : • This is a technique used mainly in immunology to

detect the presence of an antibody or an antigen in a sample.

• In ELISA an unknown amount of antigen is affixed to a surface and then a specific antibody is washed over the surface so that it can bind to the antigen.

• This antibody is linked to an enzyme and in the final step a substance is added that the enzyme can convert to some detectable signal.


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Applications in public health :– HIV test or West Nile Virus.

ELISA can be used for detecting tumour markers for certain cancers, e.g.

Prostate Specific Antigen (PSA) in carcinoma prostate.


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Fluorescent immunoassay : • When fluorescent molecules are irradiated with light at

appropriate wavelengths, an electron in the ground state is transited into the excited state. As the electron returns to the ground state, physical energy is released in the form of a photon which is detectable.

Applications in public health :• For detecting antigens in tissue sections in kidney biopsies. • Use of Auramine rhodamine stain in detection of tubercle bacilli

in smears shown to have better sensitivity than the conventional ZN stain.

• To detect the concentration of drugs, hormones and proteins and polypeptides.


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Diagnostic Technologies in Molecular Biology

Diagnostic Technologies in Molecular Biology• Molecular biology is the study of biology at a molecular

level. • Molecular biology chiefly concerns itself with

understanding the interactions between the various systems of a cell, including the interactions between DNA, RNA and protein biosynthesis and learning how these interactions are regulated.

• The tools of molecular biology is extremely useful in diagnosis, therapy, epidemiologic investigations and infection control.

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• Ease of performance, reproducibility, sensitivity and specificity of molecular tests are important, cost and potential contribution to patient care are also of concern.

• Molecular methods may be an improvement over conventional microbiologic testing as they are most practical and useful application is in detecting and identifying infectious agents for which routine growth-based culture and microscopy methods may not be adequate.

Molecular Biology contd…

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Gel electrophoresis : • It is one of the principal tools of molecular biology.

The basic principle is that DNA, RNA and proteins can all be separated by means of an electric field.

• In agarose gel electrophoresis, DNA and RNA can be separated on the basis of size by running the DNA through an agarose gel.

• Proteins can be separated on the basis of size by using an SDS-PAGE gel, or on the basis of size and their electric charge by using 2D gel electrophoresis.


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Hybridisation assays : • When the hybridization reaction is used to analyse the

nucleic acid content of an unknown sample, the process is known as a hybridization assay.

• The property of complimentary base pairing allows fragments of known composition (the probes) to interrogate an unknown for the presence of matching (complimentary) sequences.

• The detection of the hybrids can be done a variety of technologies including radioisotope labels, fluorochrome based detection and enzyme based systems.


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• Liquid phase hybridization : When both sample and probe are in solution.

• Solid support hybridization : In these assays, hybridization occurs in a biphasic environment, a solid phase (usually sample) and a liquid phase (usually probe).

• Southern and Northern hybridization assays : These combine electrophoretic separation of test nucleic acids with transfer to a solid support and subsequent hybridization.


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• The original procedure was described by EM Southern and the test nucleic acid was DNA. When RNA is the nucleic acid under test, the technique is called northern blotting by analogy.

• In situ hybridization : This is the detection of specific genetic information within a morphological context (intact tissue, cells).


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Amplification technology : • Amplification systems are enzyme based processes in

which a single enzyme or multiple enzymes synthesize copies of target nucleic acid.

• All result in production of billions of copies of the amplified product in a few hours.

• These techniques are subject to contamination from product molecules of previous amplifications and hence false positivity is high.

• However, special lab design, practices and workflow have helped to reduce false positives to an acceptable range.


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Polymerase Chain Reaction (PCR)

• It is a simple in vitro chemical reaction that permits synthesis of large quantities of nucleic acid.

• This is brought about by heating the reaction mixture to separate the strands of the DNA, then cooled to permit the primers to anneal to the target DNA in a sequence specific manner. The DNA polymerase then initiates extension from end.

• Thus the whole process is carried out in a programmable thermocycler which controls the temperatures at which various steps occur.


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• Reverse transcriptase PCR (RT-PCR) : PCR was initially described to amplify DNA. RT-PCR was developed to amplify RNA targets

• Nested PCR : This variation increases the sensitivity and specificity of PCR. The products of the first round of amplification are subject to second round of amplification with a second set of primers.

• Real time PCR : Here the target amplification and detection occur simultaneously.

• The computer software supporting the thermocycler monitors the data throughout the PCR at every cycle and generates a plot.


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Applications of molecular biology in Public Health

• Pathogen detection by hybridisation : The DNA of the infectious agent is detected directly in the clinical specimens by DNA probes.


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• DNA probes for culture identification : Probes are available for various organisms like Mycobacterium tuberculosis, Chlamydia trachomatis, Neisseria gonorrhoeae, Human papilloma virus, Hepatitis C virus, Cytomegalovirus and Herpes simplex virus.

• This is especially important for slow growing organisms like Mycobacteria, fungi etc. Identification of cultured Mycobacteria by conventional methods is slow and time consuming.

• The use of probes for this purpose permits identification from cultures within one working day and sensitivity (95.4%) and specificity (99%) are excellent.


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• DNA amplification for diagnosis : • Hepatitis C Virus- Detection of the virus by reverse

transcriptase PCR confirms current infection and has a role in diagnosis and monitoring response to therapy. Development of Quantitative PCR allows quantification of HCV which is important in prognosis and monitoring of therapy.

• HIV : Newborns upto 18 months. Quantitation of HIV virus is widely used for prognosis and for evaluation of response to antiretroviral therapy


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Mycobacterium tuberculosis :• In general the sensitivity of these assays for specimens for

which the smear for acid fast bacilli (AFB) is positive is excellent (95-100%). It is lower for specimens that are AFB smear negative (50-80%).

• Currently, the amplification processes cannot replace the AFB smear because the latter is used to determine the level of infectivity of patients and in gauging the initial response to therapy.

• At present the role of amplification assays remains complimentary to microscopy and culture.


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• Sexually Transmitted Diseases : Amplification assays have been developed for Neisseria gonorrhoeae and Chlamydia trachomatis.

• Fungi : PCR assays for diagnosis of invasive Candidiasis and Aspergillosis have been developed.

• Detection of antimicrobial resistance : Molecular detection of the Mec-A gene is now considered a reference method for assessing Methicillin resistance in Staphylococci.

• Used in detection of resistance to rifampicin and isoniazid.


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Molecular Epidemiology : • The techniques of molecular typing are useful

not only in research settings but also in real life clinical and public health problems. – e.g. prediction of response to interferon therapy is

important in patients with Hepatitis C virus infection.

• HCV genotypes 1b and 1a appear less likely to respond to interferons.


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Diagnostic Technologies in Clinical Chemistry

• Colorimetry : This is performed in a device that measures the absorbance of particular wavelengths of light by a specific solution.

• Changeable optic filters are used in the colorimeter to select the wavelength of light which the solute absorbs the most, in order to maximize accuracy.

• The usual wavelength range is from 400 to 700 nanometres (nm). In modern colorimeters the filament lamp and filters may be replaced by several light-emitting diodes of different colors.

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• Applications in public health : Routine analytes like urea, creatinine, glucose, uric acid and also hemoglobin in hematology.

• Levels in blood of routine analytes forms the basis of the diagnosis of diabetes, hyperlipidemia, metabolic syndrome etc.

Clinical Chemistry contd…

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Nephelometry : • This is a method for measuring the concentration of a solution

that contains particles that are too large for colorimetry/ absorption spectroscopy.

• When a collimated light beam strikes a particle in suspension, portions of the light are absorbed, reflected, scattered and transmitted.

• Nephelometry is the measurement of the light scattered by a particulate solution.

• Applications in public health : Measurement of antigen antibody complexes formed in immunoassays. Detection of IgG, IgM, IgA.


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Electrophoresis : Electrophoresis is the separation of charged compounds based on there electrical charge.

• Common support media include cellulose acetate, agarose and polyacrylamide gel.

• Once separation has occurred, the support medium is treated with dyes to stain and identify the separated fractions.

• To obtain a quantitative profile of the separated fractions, densitometry is performed on the stained support medium.


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Applications in public health :• Separation of serum proteins : – Patterns of hypoproteinemia in malnutrition or gross loss of

protein show decreases in all fractions with dramatic reduction in albumin.

– Cirrhosis of the liver shows a specific pattern with severe reduction of albumin and increased immunoglobulins.

– The pattern in monoclonal gammopathies (multiple myeloma) is very characteristic.

– Separation of hemoglobins : This is important in the diagnosis of various abnormal hemoglobins like thalassemia, sickle cell anemia etc.


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Chromatography : Chromatography is a separation method based on the different interaction methods of the specimen compounds with the mobile phase and with the stationary phase, as the compounds travel through a support medium. The compounds interacting more strongly with the stationary phase are retained longer in the medium than those that favor the mobile phase.

• Gas chromatography : It is used for naturally volatile or those which can be easily converted to a volatile form.

• Liquid chromatography : It is used for too unstable or insufficiently volatile for gas chromatography.

• High Performance Liquid Chromatography : It is used for analyzing air and water pollutants, monitoring pesticide levels in the environment. Used In Sickle cell Disease diagnosis in infants. Detection of additives / chemicals in food.


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• Applications in public health : Used for determination of drugs and chemicals in body fluids (toxicology).

• Detection of additives / chemicals in food by High performance liquid chromatography.

• Environmental and water pollutants can be detected with accuracy through the use of various chromatographic procedures.


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Automation in clinical chemistry : • Process a large volume of tests quickly. This is

possible through the increased speed of testing. • Most automated chemistry analyzers, photometric

methods of analysis such as colorimetry, spectrophotometry or nephelometry etc.

• Calculations, calibration curves and quality control are performed by the computers, thus reducing errors and providing more accurate results.


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• The testing pathway in an analyser may bea) Sequential testing- Multiple tests analysed one

after another on a single specimen.b) Batch testing- All samples are loaded at the same

time and a single test is conducted on each sample.

c) Parallel testing- More than one test is analysed concurrently on a given clinical specimen.

d) Random access testing- Any test can be performed on any sample in any sequence.


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Diagnostic Technologies in Hematology

Automated Instruments : • These instruments have many different components to analyze

different elements in the blood. • The cell counting components count the numbers and types of

different cells within the blood. The results are printed out or sent to a computer for review.

• It aspirate a very small amount of the specimen through narrow tubing. Within this tubing, there are sensors that count the number of cells going through it and can identify the type of cell; this is flow cytometry.

• The two main sensors used are light detectors and electrical impedance.

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• One way the instrument can tell what type of blood cell is present is by size.

• Other instruments measure different characteristics of the cells to categorize them.

• Because an automated cell counter samples and counts so many cells, the results are very precise.

• In addition to counting, measuring and analyzing red blood cells, white blood cells and platelets, automated hematology analyzers also measure the amount of hemoglobin in the blood and within each red blood cell.

Hematology contd…

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Diagnostic Technologies in Cytopathology

Exfoliative cytology : The microscopic examination of cells that have been shed from a lesion or have been recovered from a tissue for the diagnosis of disease.

Applications in public health• Cervical cytology : Exfoliative cytology from the uterine

cervix is valuable in picking up premalignant lesions of the cervix.

• It has an overall sensitivity of 80% and a specificity of 99.4% for cytologic screening for cervical cancer.

• The sensitivity was slightly lower for mild and moderate dysplasia (78.1%) and slightly higher for carcinoma in situ and severe dysplasia (81.4%) and invasive carcinoma (82.3%)

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Fine Needle Aspiration Cytology : • Fine Needle Aspiration biopsy is safer and less traumatic than an

open surgical biopsy and significant complications are usually rare, depending on the body site.

• Common complications include bruising and soreness. There is also a risk that the cells taken do not enable a definitive diagnosis.

• The technique is suitable to evaluation of lesions/ nodules arising from breast, lymph nodes, thyroid, liver, kidney and soft tissue.

• The sensitivity and specificity varies according to the organ involved and also depends on the aspiration of a representativeness of the sample.

Cytopathology contd…

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Diagnostic Technologies in Histopathology• The tissue is removed from the body and then placed

in a fixative which stabilizes the tissues to prevent autolysis.

• The samples are transferred to a cassette, a container designed to allow reagents to freely act on the tissue inside. This process is known as tissue processing.

• The processed tissue is then taken out of the cassette and set in a mold. Through this process of embedding, additional paraffin is added to create a paraffin block.


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• The process of embedding then allows the sectioning of tissues into very thin (2 – 7 micrometer) sections using a microtome.

• The microtome slices the tissue ready for microscopic examination. The slices are thinner than the average cell and are layered on a glass slide for staining.

• To see the tissue under a microscope, the sections are stained with one or more pigments.

• The most commonly used stain in histopathology is a combination of hematoxylin and eosin.


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• Applications in public health : Malignancies and also some non neoplastic conditions like tuberculosis and other infections.


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• Immunohistochemistry : Immunohistochemistry or IHC refers to the process of localizing proteins in cells of a tissue section exploiting the principle of antibodies binding specifically to antigens in biological tissues.

• Immunohistochemical staining is widely used in the diagnosis of abnormal cells such as those found in cancerous tumors.

• Applications in public health : Recognition of specific molecular markers like S100, Cytokeratin etc. helps in identifying the origin and hence the diagnosis of various tumours and conditions.


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Radiological Techniques

X-ray : For detection of pathology of the skeletal system and some disease processes in soft tissue, e.g. pneumonia, lung cancer or pulmonary edema and the abdominal X-ray, which can detect ileus (blockage of the intestine), free air (from visceral perforations) and free fluid (in ascites).

Applications in public health : Chest x ray was the backbone of National Tuberculosis Control Programme in 1962.

• However it is included in the diagnostic algorithm of RNTCP where no sputum sample is positive/ two negative smears in a symptomatic patient.

• In occupational health the ILO criterion for diagnosis of pneumoconiosis uses findings of x ray chest only.

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• DEXASCAN : DEXA Scan stands for Dual Energy X-Ray Absorptiometry. Two different types of X- ray scan the body and the images are subtracted by the computer giving a very accurate estimation of bone density/body fat.

• It is used to detect the presence of osteoporosis in men and women with particular risk factors, screen for osteoporosis, particularly in women making decisions about hormone replacement therapy at menopause.

Radiology contd…

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• Angiography : It is the study of blood vessels with a contrast medium.

• It has a great role in study of coronaries to detect the status of these vessels and decide the treatment modality for the patient.

• With the epidemic of coronary artery disease angiography will go a long way in the final management of the patients.

Radiology contd…

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Mammography : • Mammography is the process of using low-dose X-

rays, high contrast, high-resolution film and an X-ray system designed specifically for imaging the breasts.

• The goal of mammography is the early detection of breast cancer, typically through detection of characteristic masses and/or micro calcifications.

• Mammography has been shown to reduce mortality from breast cancer.

Radiology contd…

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• USG:-– Obstetric use– Cardiac scan – Renal, liver and gallbladder scans. – Musculo-skeletal imaging of muscles, ligaments

and tendons, – Eye scans – Testicle, thyroid, salivary glands and lymph nodes

scan. – To guide interventional procedures (FNAC)

Radiology contd…

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• Applications in Public Health : This technique in antenatal cases is practiced to – Date the pregnancy– Confirm fetal viability, – Determine location of fetus, – Intrauterine vs ectopic,– Location of the placenta.– Number of fetuses– Major physical abnormalities, – Assess fetal growth– Fetal movement and heartbeat.

• Ultrasound is also increasingly being used in trauma and first aid cases, with emergency ultrasound becoming a stable of most emergency response teams.

Radiology contd…

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CT scan : • CT scan works on the principle that cross sectional

slices of the body are produced using X rays, followed by processing by the computers to study detailed anatomy of the specific location or slice.

• The modification of conventional CT is spiral where the X-ray tube/ detector combination rotates continuously around the patient creating a spiral and helps in generating high quality images.

Radiology contd…

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• Magnetic Resonance Imaging : Charged spinning particles such a protons behave like tiny bar magnets and are used to produce image of biological tissue.

• It does not use ionizing radiation hence is safe with no known biological hazards.

• It provides excellent tissue contrast with good resolution.

• Images can be obtained in any plane (Multiplanar Imaging) and does not produce artifacts due to bone and is an ideal imaging modality for spine, posterior fossa and musculoskeletal system.

Radiology contd…

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Radionuclide scan : • A radionuclide scan is a way of imaging bones, organs and

other parts of the body by using a small dose of a radioactive chemical.

• Cells which are most ‘active’ in the target tissue or organ will take up more of the radionuclide. So, active parts of the tissue will emit more gamma rays than less active or inactive parts.

• The gamma rays which are emitted from inside the body are detected by the gamma camera, are converted into an electrical signal and sent to a computer.

• The computer builds a picture by converting the differing intensities of radioactivity emitted into different colours or shades of grey.

Radiology contd…

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• Public Health Importance– Bone scan : Used to detect areas of bone where

there is cancer, infection or damage.– Kidney scan : Used to detect scars on the kidney

and how well urine drains from the kidney to the bladder.

– Lung Perfusion Scan : Detects pulmonary embolus.

Radiology contd…

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CT scan, its modifications and their uses in public health

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MRI : Clinical uses

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IPHS Standards : Suggested Availability of radiological studies

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Electrophysical Technologies

• Various functional disturbances sometimes detectable only via analysis of electric potentials produced by some organs (tissues) as a manifestation of their function (ECG, EEG, EMG, ERG and others).

• Evaluation of these signals is performed either in the “time domain” (detection of particular specific peaks and wave complexes - “grapho-elements” - and description of their time and amplitude characteristics) or in the “frequency domain” (frequency spectrum characteristics).

• At present this is done almost exclusively with the use of computers.

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Electroencephalography : • This is the neurophysiologic measurement of the electrical

activity of the brain by recording from electrodes placed on the scalp or in special cases, subdurally or in the cerebral cortex. It represent an electrical signal (postsynaptic potentials) from a large number of neurons.

• Each electrode is connected to an input of a differential amplifier, which amplifies the voltage between them. The resulting voltage signal is filtered which is shown on paper or displayed on a computer screen. Uses in – Epilepsy and syncope– Sleep disorders– Coma and brain death

Electrophysiology contd…

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Electrocardiography : An electrocardiogram records the electrical activity of the heart over time.

• Electrodes on different sides of the heart measure the activity of different parts of the heart muscle.

• An ECG displays the overall rhythm of the heart and weaknesses in different parts of the heart muscle.

• Applications in public health : It is the best way to measure and diagnose abnormal rhythms of the heart in– Damage to the conductive tissue carrying electrical signals, – High or low potassium. – Myocardial infarction (MI)


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Electromyography : • Electromyography (EMG) is a technique for

evaluating and recording the activation signal of muscles.

• An electromyograph detects the electrical potential generated by muscle cells when these cells contract and also when the cells are at rest.

• EMG is used to diagnose two general categories of disease : neuropathies and myopathies.


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ENDOSCopy

• Different types of endoscopies are performed for diagnostic purposes of carcinoma and other pathologies. These may be rigid or fibreoptic.

• Direct:-– Laryngoscopy– Broncoscopy– Colonoscopy– Gastroscopy– Nasopharyngoscopy– Sigmoidoscopy– Hysteroscopy• Indirect:-– Laryngoscopy

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References

• Rajvir Balwar, Text book Public Health and Community Medicine, 1st Edition, AFMC Pune in collabration with WHO, Dept Of Commnity Medicine, 2008.

• Indian Public Health Standards (IPHS) Guidelines for Sub-District/Sub-Divisional Hospitals (31 to 100 Bedded) Revised 2012 , DGHS, MOHFW, Govt Of India, June 2012.

• Indian Public Health Standards (IPHS) Guidelines for District Hospitals (101 to 500 Bedded) Revised 2012, DGHS, MOHFW, Govt Of India, June 2012.

• http://www.biomerieux.com/en/advanced-technologies• World Health Organization, Laboratory Support to Emerging Diseases. World

Health Organization Regional Office for South-East Asia New Delhi, 2008.• www.whoindia.org/LinkFiles/

Public_Health_Laboratory_Networking_SEARO_recommendations _on_networking_of_PHL

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• The Diagnostics Innovation Map: Medical Diagnostics for the Unmet Needs of the Developing World, WHO BIO Ventures for Global Health,2010.

• Estimating the Global Health Impact of Improved Diagnostic Tools for the Developing World. Santa Monica, Rand Institute,2007.

• Sen J, Belli A "S100B in neuropathologic states: the CRP of the brain?". J. Neurosci. Res.2007; 85 (7): 1373–80.

• http://mohfw.nic.in/NRHM/iphs.htm• http://testequipment.globalspec.com/LearnMore/

Labware_Scientific_Instruments/Analytical_Instruments/Chromatographs/HPLC

• http://www.google.co.in/imgres?um/diagnostic%technologies/php.

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