thinking like a scientist

Question 1

Accuracy and Precision

Answer 1

Accuracy is a measure of how close a single measurement from a set is to the accepted true value.

Precision measures the closeness with which repeated analyses of given samples agree with each other. It is usually expressed as a standard deviation, which is a quantitative measure of the spread of a set of measurements.

Question 2

Accuracy and Precision

Answer 2

Accuracy is a measure of how close a single measurement from a set is to the accepted true value.

Precision measures the closeness with which repeated analyses of given samples agree with each other. It is usually expressed as a standard deviation, which is a quantitative measure of the spread of a set of measurements

Question 3

Screening and diagnosis

Answer 3

SCREENING TEST :
used to identify those individuals who are a high enough risk (in a population) to warrant diagnostic processes which are too expensive to offer everyone

Screening is also offered where a specific population may be at risk of a particular disease - subgroup of the population who is at a greater risk of a disease has been identified

diagnosis:
is facilitated using a more accurate test or group of tests or procedures. These diagnostic tests will confirm whether an individual has that disease or not.

Question 4

Qualitative and quantitative tests

Answer 4

QUALITATIVE TEST :
identifying what is present in a sample, or determining whether a particular compound is present or not.

This is called a qualitative test, which can be defined as “a test which is concerned with only the nature of the substance under investigation and not its amount or concentration”

In qualitative tests colour changes or a visual clue is often used to indicate whether a substance is present or not.

QUANTITATIVE TEST : which is “concerned with the amount or concentration of the substance(s) under investigation”
An example is the pregnancy test kit which detects the presence of a hormone in urine.
REMEMBER :
It is always important to quote the units of a value when reporting quantitative results. The international system of SI units is frequently used, but not always. In many applications, many different units are used, depending on the context

Question 5

QUALITATIVE ANALYSIS :

a presumptive test

a confirmatory test

Answer 5

A presumptive test is a qualitative analysis that confirms that (a) the sample is definitely NOT a specific thing, or (b) that a sample is probably a specific thing.

A confirmatory test is one that confirms the original finding. tend to cost more

Question 6

Selectivity, specificity and sensitivity

Answer 6

Selectivity:
the ability to discriminate between the target analyte and other constituents of the sample which may interfere with the analysis

Specificity means an analytical method is perfectly selective for an analyte or a group of analytes

Sensitivity is defined as the ability to discriminate between small differences in the concentration of an analyte in a sample, or the ability to detect (qualitative analysis) or determine (quantitative analysis) very small amounts of analyte

Question 7

An analyte

Answer 7

An analyte is the substance or component you want to analyse.
For example, if an analytical method can distinguish one molecule in a mixture of similar molecules, it is selective for that molecule

The more similar the concentrations of analyte that can be distinguished, or the lower the amount of analyte that can be detected or determined, the higher the sensitivity of the technique.

Question 8

false positives

false negatives

Answer 8

false positive:
the analyst needs to know how many times the test returns a positive result when the sample is actually negative

a false negative:
sometimes the test will produce a negative result when the sample is actually positive
The terms ‘sensitivity’ and ‘specificity’ are used to describe this too;

Question 9

standard and calibration

Answer 9

standard, which is “an analyte of known concentration”

Calibration is a set of operations that establishes, under specified conditions, the relationship between an instrument response and the corresponding values of a standard

Question 10

doing a calibration

Answer 10

Calibration can be done in many ways but a common method is the external calibration procedure. This is done by using several chemical standards made up to a known concentration across the concentration range of interest. Ideally, more than one standard is made up for each concentration to obtain replicate values.
A calibration profile by convention is plotted with the instrument reading for each concentration on the vertical (y) axis and the standard concentrations on the horizontal (x) axis. The calibration standards are measured in the analytical instrument under the same conditions as those subsequently used for the unknown samples. The analyte concentrations in the real samples are then obtained from the calibration graph.

Question 11

Limit of detection (LOD) –

Answer 11

this is the lowest quantity of a substance that can be distinguished from the absence of that substance (a blank value) within a stated confidence limit (generally 1%).

Question 12

Limit of quantification (LOQ) .

Answer 12

this is the lowest concentration that can be identified and quantitatively measured using an analytical method that has been validated with specified accuracy and precision. The two differ in that, while the LOD is the ultimate lower limit, the LOQ considers potential ‘noise’ in the system (potentially generated by errors in measuring).

Question 13

Limit of linearity (LOL) –

Answer 13

this is the concentration at which the calibration curve departs from linearity by a specified amount. A deviation of approximately 5% is usually considered the upper limit. This is common at higher concentrations. After this point, the relationship between generated signal and concentration is no longer predictable.

Question 14

health and safety

Answer 14

There are scientific principles underlying safe practice procedures and the equipment used in any professional practice

able to identify sources of hazard in the workplace and to make evidence-based judgements about whether an intervention is potentially more beneficial than harmful. It is important to consider how to minimise risks by selecting the appropriate strategies for hazard control and risk management.

Question 15

hazard

Answer 15

A hazard is defined as the source of something that may cause harm, damage or adverse health effects; risk is the probability that someone may be harmed after being exposed to a hazard

Question 16

Workplace activities in the UK are subject to health and safety legislation. They are covered under Section 7 of the Health and Safety at Work, etc. Act 1974 which states that:

Answer 16

It is the duty of every employer, so far as is reasonably practicable, to ensure the health, safety and welfare at work of all employees.

It is the duty of every employee while at work to take reasonable care of himself (or herself) and of other persons who may be affected by his (or her) acts or omissions at work.

The employees must cooperate with the employer with regard to health and safety procedures.

Question 17

personal protective equipment (PPE)

regulations, the Control of Substances Hazardous to Health (COSHH)

Regulations and the Reporting of Injuries, Diseases and Dangerous Occurrences Regulations (RIDDOR)

Answer 17

The essential requirements of COSHH are that the employer must:
• Assess the health risk to employees;
• prevent or control the exposure of employees to hazardous substances.

Question 18

Whether the work is done in a laboratory or in the field, the analytical scientist is protected under this legislation, provided he or she follows some basic dos and don’ts:

Answer 18

Laboratory coats or protective clothing must be worn at all times.

When handling chemicals, biological samples or ‘sharps’ (i.e. any sharp object that can cause injury, particularly to the hands), observe good laboratory practice by wearing gloves. Latex gloves were widely used but are less common now because of the risk of serious allergic reactions. Nitrile gloves are now the most widely used, depending on the application.

There should be no eating, chewing of gum, drinking, smoking or applying of cosmetics in any laboratory when sample collecting, preparation or analysis is under way.

Do not pipette (draw up a liquid by suction) by mouth; always use a pipette filler.

All broken glass must be placed in a labelled bin

Question 19

biological hazards

chemical hazards

Physical hazards

radioactive substances

Answer 19

biological hazards
blood, urine, faeces, dangerous pathogens (a pathogen is a disease-causing organism).

chemical hazards are toxic, corrosive or harmful chemicals which may be inhaled, ingested or come into contact with skin.

Physical hazards include high voltages from some analytical instruments or even the normal household electricity supply, which can deliver a fatal current.

Each organisation using radioactive sources will probably have local rules about the procedures for ordering, storage, handling, disposal and record keeping.
Radiation laboratories are controlled areas with specific access requirements. All radioactive material must be accounted for as the loss of material can result in prosecution. Secure containment is required by law, to prevent reagents being stolen, and to protect everyone working in the laboratories

Question 20

QUALITY

Answer 20

Quality is an important concept in the standardisation of methods, i.e. ensuring methods work according to a particular defined standard or method.

quality can be defined as the body of characteristics, properties, attributes or abilities of an entity (an object, a system or a service) that make it better or worse than, or equal to, other entities of the same type

The International Organisation for Standardisation (ISO) defines quality as ‘the totality of characteristics of an entity that bear on its ability to satisfy stated and implicit needs imposed by a client or through legislation’. Consequently, quality also involves usefulness, fulfilling requirements and satisfying clients’ needs
In terms of the results obtained in the analytical laboratory, quality does not necessarily mean getting the most accurate results; quality in this context means matching the service with the requirements of the client. This is commonly described as results that are ‘fit for purpose’.

Question 21

METHOD

Answer 21

A method is a set process that if followed will ensure accurate and precise results.

Question 22

Quality in the analytical laboratory is achieved by providing analytical results that:

Answer 22

• meet the specific needs of the client
• attract the confidence of the client
• represent good value for money.

Question 23

Quality assurance (QA)

Answer 23

describes the overall measures that an analytical laboratory uses to ensure the quality of its service. It focuses on providing confidence that quality requirements will be fulfilled and reliable analytical measurements will be produced.

Including staff training, record keeping, ensuring a suitable laboratory environment (such as adequate storage facilities to ensure the integrity of samples or filtered air conditioning to stabilise temperature and humidity and reduce dust levels), maintenance and calibration schedules
for instruments, the use of validated and documented methods, and preventive and corrective actions.

Question 24

Quality control (QC

Answer 24

involves examining the laboratory and its analytical results. QC encompasses the planned activities designed to verify the quality of the measurement and ensure that the analytical processes are working correctly

There are two types of QC:

• internal QC, which provides confidence to the laboratory staff carrying out the measurements

• external QC, also known as external quality assessment, which provides confidence to the client by comparing the performance of the laboratory with others making similar
measurements.

Although serious flaws in laboratory processes are sometimes highlighted, they are comparatively rare considering the number of tests that are carried out in laboratories every year.

Question 25

ISO (International Organisation for Standardisation)

Answer 25

ISO enable laboratories to take an ‘off-the-shelf’ standard that best suits their organisation and design a quality management system that meets the standard’s requirements

here are numerous standards but the common ones relevant to laboratories are as follows:

BS EN ISO 9001:2008 Quality management systems – Requirements relates primarily to promoting the effectiveness of quality management and enhancing customer satisfaction (BSI, 2008). This standard is a very general document and does not cover any technical aspects.

BS EN ISO/IEC 17025:2005 General requirements for the competence of testing and calibration laboratories addresses the technical competence of laboratories to carry out specific tests and calibrations (BSI, 2005). It is used by accreditation bodies worldwide as the core requirement for the accreditation of laboratories.

Question 26

certified laboratory and an accredited laboratory.

Answer 26

Certification represents a written assurance by an independent third party of the conformity of a product, process or service to specified requirements.
Does not specifically address technical competence.

accreditation is the formal recognition by an authoritative body of competence to carry out specific tasks to specified standards.

Question 27

GLP

Answer 27

The Organisation for Economic Co-operation and Development (OECD) has developed the Good Laboratory Practice principles, commonly known as GLP:
GLP is a legal requirement for all laboratories doing regulatory work, ensuring the production of quality test data. It embodies a set of principles that provides a framework within which laboratory studies are planned, carried out, monitored, recorded, reported and archived.

Question 28

Good Manufacturing Practice (commonly known as GMP).

Answer 28

GMP has been defined as the part of quality assurance that is aimed at ensuring that products (such as food, pharmaceuticals and medical devices) are consistently manufactured to a quality appropriate to their intended use. In the UK, the Medicines and Healthcare products Regulatory Agency (MHRA) enforces GMPs.

Question 29

The Scientific Method

Answer 29

each stage of the process has a significant bearing on those that come before or after and, in some instances, not all of the steps are required.

Define the problem
(consider possible experimental procedures)

Devise sampling strategy and collect samples

Sample preparation and carrying out the analysis

Data analysis
(including statistical analysis)

Interpret data and statistics in relation to analytical problem

Draw conclusions and/or recommend new analyses

The method by which a sample is obtained can influence whether or not results are reproducible; however, sampling may be constrained, or done in a particular way, because the analytical process is driven by a particular problem.

Question 30

Defining the problem:

A hypothesis

Answer 30

hypothesis is a testable scientific idea or problem that can be used to make predictions of behaviour. It suggests an answer to a problem or an explanation for an observation, which can then be tested analytically and statistically, and generally states a relationship between two or more variables. In essence it is a question based on the initial observations.

Once a hypothesis is devised, a set of focused research questions can be framed around it, based on what is already known and what gaps exist on our knowledge.

Question 31

A Theory

Answer 31

Once a hypothesis has been tested repeatedly, and experiments show the predicted behaviour, it can be developed into a theory. If a theory is disproved through further analysis then a new theory needs to be developed to refine or replace the old one.

The cycle of setting up a hypothesis, testing it, refining or reformulating it, retesting it, and so on, is often called the scientific method and it underpins all scientific investigations – analytical or otherwise.

Question 32

BULK SAMPLE

Answer 32

bulk sample – sometimes called gross samples – represent the sample before it is divided into aliquots.

An aliquot is also known as sub-sample.
Aliquots separated from the bulk can be used for several analytical techniques and/or for replicates of a single analysis

Question 33

REPLICATE

Answer 33

A replicate is a repeat experiment that can be used to ensure your test is accurate and precise.

Question 34

The most important initial consideration is to establish whether the data required should be qualitative or quantitative.

Answer 34

Once this information is established, other analytical considerations are needed:
• How accurate does the analysis need to be?
• How much sample is available versus how much sample is needed?
• Does the sample need to be manipulated (extracted, stabilised, etc.) before analysis?
• What is the likely concentration range(s) of the analyte(s)?
• How sensitive is the technique? What is the limit of detection (LOD) and the limit of
quantification (LOQ)? Can it measure what you need?
The analytical considerations listed above are intrinsic to the analysis, sample, hypothesis and problem

Question 35

other factors that can also affect the selection of a suitable technique for pharmaceutical analysis. These are:

Answer 35

Speed – Data may be required quickly, or there may be numerous samples that would occupy an analyst or instrument for a long time

• Skills required of analyst – training may be needed, or the assistance of a trained operator.
• Cost of equipment and infrastructure – not every laboratory can afford to provide every possible analytical instrument or facility. Commercial laboratories may contract other laboratories to complete analyses using otherwise unavailable techniques
• Availability of equipment –The ideal analytical instrument may not be available when the analyst needs it, in which case an alternative must be sought.
• Per-sample cost – If the cost per sample is too high, a less expensive, alternative technique may be chosen.
• Health and safety – Some analytical techniques require the use of biological materials,that may be hazardous to the analyst.

Question 36

peer review

Answer 36

Peer review is the process by which the scientific community self-regulates itself by requiring scientists to assess the work of other scientists and provide an independent, unbiased and critical assessment. QA and QC is key to the validity of results. When QA is done in pharmaceutical research laboratories, the practice is highly specific and rigorous

Question 37

data analysis

Answer 37

the reduction of data (otherwise known as data analysis) and the application of that data to the problem (known as data interpretation). Data analysis and interpretation are, generally, approached by the use of statistics.

The ultimate aim of collecting data is usually not simply to describe but also to answer scientific questions as objectively as possible.
statistical techniques offer ways of dealing with even small-scale variability.
Your data and interpretation are key.
An incorrect interpretation can have a financial, an environmental, legal or even a political impact.

Question 38

The final step in the analytical process is to formulate a conclusion.

Answer 38

all other considerations are tied together to determine whether a hypothesis has been proven or not. You should realise that even if a hypothesis is proved to be incorrect, that is still a valid and important result.

reviewing the data generated and its accompanying statistics, assessing the validity of the analytical method, and identifying any aspect that was flawed or could be improved.
Often the conclusions may inevitably lead to further questions that require additional analysis using the same technique, different samples, or an entirely new analytical process