new gen diagnostics Flashcards
Next generation DNA sequencing (NGS)
a number of different modern sequencing technologies
Examples of next generation DNA sequencing
llumina (Solexa) sequencin
Roche 454 sequencing
Ion torrent: Proton / PGM seque
ncing
SOLiD sequencing
Purpose of NGS
These recent technologies allow us to sequence DNA and RNA much more quickly and cheaply than the previously used Sanger sequencing, and as such have revolutionised the study of genomics and molecular biology.
Whole exome sequencing (WES)
massively parallel NGS of exonic sequences
Purpose of WES
WES has been enormously successful in the identification of novel Mendelian disease associated genes, and it has recently begun to be used for clinical diagnostics.
Personalised medicine
detect and eliminate tumors before they become life threatening–before they are even visible on medical images, get diagnostics to the most sensitive level possible
Next generation diagnostics functions
In general,
- Very predictive and allegedly accurate
- Lowers the risk of developing diseases due to being
- Able to actively seek out preventive measures
- Knowledge from the genome data collected can contribute to genome mapping and trace the history and development of pathogens
- Helps to control disease outbreak
Next generation diagnostics
Next-generation sequencing and personalized medicine
the shift of the emphasis of care
Prevention -> Early detection of disease -> Management of multiple chronic conditions
With current trends and time, the emphasis of care is shifting toward prevention and early detection of disease, as well as management of multiple chronic conditions. instead of focusing on finding cures. We only do so when endpoint of diseases are reached.
Point of care (POC) diagnostics
Defined as tests that are carried out near site of patient care AND outside a centralized laboratory
The 4 key requirements of POC
- simple to use
- reagents and consumables are almost in storage and usage
- device together with associated reagents and consumables are safe to use
- results should be concordant with an established laboratory method
concordant - in agreement with
Goal of POC
minimise the time to obtain a test result, allowing clinicians and patients to make a quick clinical decision
gives immediate results in non-laboratory settings to support more patient-centered approaches to healthcare delivery.
allows patient diagnoses in the physician’s office, an ambulance, the home, the field, or in the hospital. The results of care are timely, and allow rapid treatment to the patient
Outcomes of the goals of POC
Facilitate immediate evidence-based medical decisions that improves patients’ outcomes
Reduce patient acuity, criticality, morbidity and mortality especially during life-threatening crises and emergency resuscitations
Guidelines issues for PoCT devices assured guidelines provided by WHO
Assured
Affordable – for those at risk of infection
Sensitive – minimal false negatives
Specific – minimal false positives
User-friendly – simple to perform, uses non-invasive specimens
Rapid & Robust – short turnaround time and no need for refrigerated storage
Equipment-free – no complex equipment
Exceptions to assured guidelines
However, GUIDELINES for those developing PoCT devices for the detection of sexually transmitted infections (STI) provided by WHO may defer:
For PoCT devices that will be used in the developed world, some of the ASSURED criteria will also remain relevant but others will be substantially different. Thus instead of being equipment free, the need will be for relatively sophisticated equipment that at a minimum can provide a quantitative result, presentation of the results, decision support and, ideally, connectivity to other information systems such as the patient’s electronic health record. While the technology to provide all these features undoubtedly exists, they come at a cost which may be difficult to recover using the most common business model for PoCT which is that used for the central laboratory, based on complexity and reagent costs, thus only charging for the test strips/cartridges. When one combines these equipment needs with what are seen as other competitive requirements such as a small sample volume, whole blood, production of a result within 10 min of applying the sample, ease of use and requisite analytical performance, it is possible to appreciate the technological challenges involved in building such devices.
PocT market
over-the-counter products such as glucose monitoring and pregnancy testing
and
professional market includes all other testing including critical care, infectious disease, cardiac markers, diabetes, lipids, coagulation and haemotology
Basic dipsticks as with urinalysis
over-the-counter POC
a urine sample is applied to a porous pad containing reagents and a reflectance technology is used to provide a semi-quantitative estimate of the analyte
Handheld devices like glucose meters
over-the-counter POC
get a drop of blood and apply the blood to the test strip when the blood symbol appears in the meter window
the meter will count down and display the result of blood glucose
Rapid diagnostic tests
examples of POC diagnostics
Rapid antibody tests
- rapid HIV test
- rapid plasma reagin
Rapid antigen test
- rapid influenza diagnostic test
- malaria antigen detection test
- rapid strep test
Spartan RX platform
to detect mutations in the CYP2C19 gene, which codes for the CYP2C19 protein in the body.
DNA testing
Mutations in the CYP2C19 gene cause reduced response to clopidogrel, an anti-platelet aggregation drug given to patients undergoing percutaneous coronary interventions (PCI), resulting in adverse consequences
PCIs are often performed urgently, hence the potential benefits for rapid PoCT to detect mutations in the CYP2C19 gene and alternative drug therapy with better outcomes.
The desktop instruments can extract and analyze DNA from a buccal swab. It is then placed in a cartridge and inserted into the instrument and results come out in 1 hour.
There is limited data on the outcomes of using the test but the test was useful in identifying patients at risk of a poor response to clopidrogrel.
POC diagnostics in Singapore
BiosensoriX sells a variety of POC detection products for research use only
ELLI tm, Stack Pad tm, Quiz PCR Biochip Test
ELLi tm
a POC test for research use only
ELLI™ is a USB-like device that enables accurate quantification of biomarkers and pathogens for a range of antibody-based assays
- target applications that requires test and accurate biomarkers measurements
- dengue triage
- stroke triage
- acute kidney injury
Features of ELLi tm
- Easy multiplexing (multiple signals are combined into one signal over a shared medium)
- One step procedure
- High sensitivity
- Accessible to everyone
Cell ID Singapore
Portable genetic test for Covid-19
Using a nasal swab or saliva specimen, this portable genetic test uses rapid molecular testing to determine in minutes, anywhere, anytime, if a person has COVID-19 or not
They detect the genetic information of the virus, the RNA
Features of Cell ID Singapore Quiz PCR BiochipTest
Nasal Swab
One step procedure
Immediate Results
Contact Trace
Emerging POC Diagnostics function
- Blood Glucose Testings (effective options for frequent and timely measurements of blood glucose)
- Cancer biomarker (simplifying cancer biomarker test)
- HIV testing (increase the coverage of testing and expand the scope of this technology to the rural populace)
- Portable PCR test for COVID-19 (test to be sued outside of a laboratory setting and by healthcare professionals on the pandemic front lines)
Future POC diagnostics
Microbiology
- outbreaks
- methicillin resistant staph. aureus
Endocrine testing to guide surgical therapy Para-thyroid hormones - ACTH (adrenocorticotropic hormone) - Gastrin - Growth Hormones
Sepsis Markers
Stroke Markers
DNA Testing
background of ACTH
a polypeptide tropic hormone produced and secreted by the anterior pituitary gland. It is also used as a medication and diagnostic agent
Why think about ACTH ?
Deficiency of ACTH is a sign of secondary adrenal insufficiency (suppressed production of ACTH due to an impairment of the pituitary gland or hypothalamus, cf. hypopituitarism) or tertiary adrenal insufficiency (disease of the hypothalamus, with a decrease in the release of corticotropin releasing hormone CRH).
Conversely, chronically elevated ACTH levels occur in primary adrenal insufficiency (e.g. Addison’s disease) when adrenal gland production of cortisol is chronically deficient.
In Cushing’s disease a pituitary tumor is the cause of elevated ACTH (from the anterior pituitary) and an excess of cortisol (hypercortisolism) – this constellation of signs and symptoms is known as Cushing’s syndrome.
Future POC Diagnostics - AI Technologies
- Detecting Diseases - AI-based software can tell if a patient has a certain disease even before evident symptom appears
- Classifying diseases - accurately spot signs of a certain disease in medical images such as MRIs, X-rays, and CT scans
- Improving decision making process - help doctors to overcome research obstacles, process vast amounts of health data quickly, and ensure holistic understanding of a patient health
- AI-based treatment solutions - improve surgical robots that execute highly complex operations
- Making people live longer - help track immunosenescence levels and identify new interventions designed to boost the immune system
Multiple POC testing (xPOCT)
- the stimulatenous on-site detection of different analytics from a single specimen
- gathered increasing importance for clinical diagnostics with emerging applications in resource-based settings
- in addition, it is highly desirable to screen various analytes simultaneously, enabling a rapid, low-cost and reliable quantification
- However, in many instances, clinical evidence based on a single biomarker is not sufficient for an adequate diagnosis of a disease and monitoring of a treatment
Multiplexed-point-of-care testing (xPOCT)
simultaneous on-site detection of different analytes from a single specimen, has recently gained increasing importance for clinical diagnostics, with emerging applications in resource-limited settings (such as in the developing world, in doctors’ offices, or directly at home).
clinical xPOCT applications available
many different commercial devices for the simultaneous detection of clinical chemistry parameters, including blood gases, and electrolytes, or acute metabolites such as Abbott i-STAT system, Abaxis Piccolo Xpress, or Nova Biomedical StatSensor), or immunoassays (e.g., Radiometer AQT90 Radiometer and Mitsubishi PATHFAST analyzer
These systems are either bulky and expensive bench-top bioanalyzer, or only capable of detecting a limited amount of type of analytes
the future of xPOCT device will be driven by
- novel biotechnologies
- targets (e.g circulating RNAs, or tumor cells, exosomes and miRNAs)
- applications like personalized medicine, homecare monitoring and wearable
Requirements of xPOCT devices
- easily accessible
- user-friendly
- rapid-turnaround times
- longer shelf life
- accurate and quantitative results
- low-cost and portable devices
- Hi-technology devices
easily accessible - low sample consumptions or possibility to use easily accessible or noninvasive to obtain a maximum amount of information from valuable specimens
user-friendly - simple or automated system operation requiring minimal user interaction
rapid turnaround times - allow an intermediate treatment
longer shelf life - prolonged reagent storage and shelf life
accurate and quantitative results - in accordance with clinical and central laboratory findings, adhering to international quality standards
low-cost and portable devices - equipped with disposable and cartridges or strips
Hi-technology - equipment-free or cell-phone-based systems are highly-preferred
full requirements of xPOCT diagnostic devices
(i) low sample consumption (e.g., blood from a finger prick), or the possibility to use easily accessible (noninvasive) samples, like urine, saliva, sweat and breath condensate, along with a maximum amount of information obtained from valuable specimens
(ii) simple or automated system operation requiring minimal user intervention
(iii) rapid turnaround times – within 10 minutes to 2 hours – allowing an immediate treatment
(iv) prolonged reagent storage and shelf life
(v) accurate and quantitative results in accordance with clinical and central laboratory findings, adhering to international quality standards (ISO 15189);
(vi) low-cost and portable readout devices, equipped with disposable test cartridges or strips, satisfying the in vitro diagnostics guidelines (EU Directive 98/79/EC or FDA regulations). Finally, equipment-free or cell-phone-based systems are highly preferred in the developing world or other resource-poor settings [4, 5]. Ideally, the xPOCT device should also be able to analyze different types of compounds simultaneously, e.g., RNAs, metabolites, proteins, exosomes, and cells.
ideal xPOCT device
should also be able to analyze different types of compounds simultaneously
microfluidic paper-based analytical devices - μPADs
low cost - Low-cost xPOCT solutions have attracted particular interest in the developing world, e.g., for hepatotoxicity screening.
side effects - Low-cost xPOCT solutions have attracted particular interest in the developing world, e.g., for hepatotoxicity screening.
liver function testing - On-site testing by gauging the levels of two liver enzymes in whole blood, aspartate aminotransferase and alanine aminotransferase
colorimetric readout - It offers results with both qualitative and quantitative data analysis within 15 min after the sample introduction (< 35 μl blood
