Acknowledgements

1
Acknowledgements
Greg Applegate Roy Sumner John Kane Eric
Gowland Lesley Tetlow Dianne Bamber Ian
Ward Aveline Hume Sue Thornton Liz
Stewart Colin Beardwell Julia Anderson Ken
Sowerby Gilbert Wieringa Dinah Jagger Jean
Denham Sheila Gowans Janet Yates Lesley
Potton Kath Hayden Rachel Haggas Kath
Brownbill Angela Rees Helen Sumner Kirstie
farrell Annal Rowbottom Pete Smeaton Anne
Ryan Kim Rowbottom Celia Yates Adoption
Pravin Patel (Physics) Roger Burrell(from
LKB) Robert Moore (Quattro) Hovering
above Yalow Berson Ekins Priceless
animals Edgar (Sheep) Edward
(Donkey)
2
Immunoassay ?Navel Gazing
(Radio)immunoassay is the most important
development in medicine since penicillin (in the
developed world). Discuss
elr 1983
3
Health and Safety
Radiation Exposure risk in context Protection
measures/monitoring Excuse to go to the
pub! Radiolabelling (hot hair/shoes) Hormones
Making up oestradiol standards
4
Assay Design
Theoretical Knowledge Saturation analysis,
antigen, antibody structure, hapten, ligand,
conjugates, adjuvant, titres, molecular
structures, molecular weights (cold room door),
immunogenicity, affinity constants(Ka), reaction
kinetics, radioactivity, half-lives, hook effect,
clinical endocrinology etc Sample
storage Immediate separation Store frozen or
assay same day Pipetting Reverse
pipetting Extensive training Dilutions Regular
checks of accuracy and precision
5
Assay Design (contd
Incubation times Very low density of
molecules Long incubation times Attempts to
shorten the times Radiation Counting Magical
10,000 counts TRT/Additional counters Ekins
Assay Sensitivity Definition Slope(YB) vs
MLD(Ekins) Multiple zeros
6
Assay Design (contd)
Assay Specificity Parallel interferent
curve T3/T4 Curve Fitting Procedures Simple
Dose vs Response! Not Immunoassayists 1.
Change counts to binding 2. Subtract
NSB from B 3. Log transform the
concentration 4. 10 ways to join
points!
7
Assay Design (contd)
Assessment of Linearity (Parallelism) Dilution
of patient sample(s) Matrix effects c.f.
recovery Free Hormone Measurement Rationale Pra
cticality vs Purism (Ekins)
8
Quality Assessment
Internal QC 1. Between batch variation 2.
Within batch variation 3. Maximum Binding 4.
Non-specific Binding 5. Slope parameters 6. Fit
parameters 7. Assay means etc (These
figures were examined then stored for years - A
CPA Inspectors dream.)
9
Quality Assessment (contd)
External QA Extras 1. EQAS is mainly for
bias. 2.Comparison to true result. 3.
Recoveries. 4. Tests of base-line securit.y 5.
Tests for Specificity. 6. Tests for data
handling (sent raw counts). 7. Tests of Clinical
Interpretation (20 years BC). 8. Better liaison
with participants. Precision profiles A measure
of performance throughout the range Often used
to determine useful assay range
10
Reporting
Clinical Authorisation Manual delta
checking Duplicates SDAMs MOMs Results
pattern recognition Cumulative
reporting Discussion with clinicians Adding
Comments Rationale Etiquette Computerisation I
s it of value?
11
Additional Navels (to gaze at)
Standardisation Units Reference
ranges Which thyroid hormones?
(CS) Replicates Macromolecules Monoclonal
antibodies Alternative labels Automation
etc etc
12
Conclusions

• Was it all a waste of time and resources?
• Why this concentration on minutiae?
• - a million times the sensitivity?
• - biological technique?
• - time to think during long incubations?
• - assay disasters were common?
• - the personalities involved?

13
The End
Thank-you L Good-Bye J Good Luck