1. Biological Evaluation Lecture - 3 Principles and Importance of BA’s Joseph O. Oweta B. Pharm (MUST)

2. Characteristics of A good Bioassay Sensitivity: Also called the true positive rate, measures the proportion of actual positives which are correctly identified as such (e.g. the percentage of sick people who are correctly identified as having the condition). Specificity Sometimes called the true negative rate, measures the proportion of negatives which are correctly identified as such (e.g. the percentage of healthy people who are correctly identified as not having the condition). A perfect assay would be described as 100% sensitive (i.e. predicting all people from the sick group as sick) and 100% specific (i.e. not predicting anyone from the healthy group as sick)

3. Characteristics of A good Bioassay Repeatability Repeatability or test–retest reliability is the variation in measurements taken by a single person or instrument on the same item and under the same conditions. A less-than-perfect test–retest reliability causes test–retest variability Reproducibility Reproducibility is the ability of an entire assay or study to be reproduced, either by the researcher or by someone else working independently.

4. Characteristics of A good Bioassay Accuracy Accuracy of a measurement system is the degree of closeness of measurements of a quantity to that quantity's actual (true) value.. Precision The precision of a measurement system is the degree to which repeated measurements under unchanged conditions show the same results

5. Characteristics of A good Bioassay Low accuracy: poor trueness, good precision

6. Characteristics of A good Bioassay Low accuracy: good trueness, poor precision

7. Note Bioassays may be performed: 1.In Vivo (“Within the living”) Using an intact animal 1.In Vitro (“Within Glass”) – Isolated Tissues – Specific Cells – Organisms in Culture

8. Advantages of isolated tissue experiments over intact animal experiments Several preparations can be obtained form a single animal Relatively small amount of test material (reagents etc.) is required Cheap, Less time consuming Interferences due to pharmacokinetic and Compensatory Reflexes is avoided

9. Choice of Tissue Should give measurable responses with discrimination in the practical range of concentrations. Should be able to differentiate between compounds.. e.g. antibiotics? Tissue should behave similar to human tissue with respect to the receptor and drug response Availability and Feasibility of test animals (Rationalisation!!) Time consumption – Dosing Cycle – Agonist Vs. Antagonist being assayed?!

10. Disadvantages of Bioassays Biological assays leave much to be desired in several respects. 1.Although some are extremely sensitive in detecting small differences in concentration, their quantitative accuracy usually falls considerably below that obtainable with chemical Techniques,

11. Disadvantages of Bioassays The techniques and interpretations involved often can vary with different operators, in spite of the rigid requirements, specified by official publications; Hence there is a considerable subjective element present.

12. Disadvantages of Bioassays The effect measured in the test animals often is not that which the drug is intended to produce in treating patients The importance of this discrepancy was minimised formerly, but recent studies have shown that when several active principles are present in a crude drug, those producing the maximal therapeutic effect are not necessarily the ones chiefly responsible for the action measured in the assay. As a result, samples found to be of equal strength by assay may show different potencies when employed clinincally. Assignment: Read and Make notes about Animal Assay of Digitaloid Drugs (David B. T. (2000) Remington: The science and practice of pharmacy. 21 st Ed. Pp. 553 – 565)

13. Classification of Bioassays Bioassays are classified in three groups according to whether: – The effect produced is all or none (death) – Graded (as rise in blood pressure) – Characterised by developing in a measured period of time (Curative response of thiamine) NB 1.In all three types, with a few exceptions, the calculations of potency are based on the sizes of doses necessary to produce approximately equal effects and not on the intensities of the responses. 2.The results derived from all are quantitative in that the potency of the unknown is expressed in terms of the standard.

14. Types of Bioassay 1.Quantal (Direct end point) Assay 2.Graded Assay 1.Bracketing Assay 2.Matching Assay 3.Interpolation assay 4.Multiple point assay

15. Quantal response (all or none) assay In this assay, the dose of standard and Unknown which provide predetermined “all or none” response are administered Their potency ratios are then compared E.g. – Digitalis induced cardiac arrest in cats – Insulin induced hypoglycaemic convulsions in mice.

16. Quantal response (all or none) assay An extract containing digitalis is infused into the vein of a cat until the heart stops. The volume of the fluid infused is noted as the threshold volume TD50 for a given test and standard calculated and compared

17. Quantal response (all or none) assay Pros and Cons AdvantagesDisadvantages Drug effects appear rapidly and easily recognised Only toxicity or high dose study possible Effect directly proportional to the drug Dose Dose Ranging can’t be done Rapid End Point detection

18. Any Questions ….next time we shall discuss Graded response assays