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Hi Katherine, I agree completely. The gold standard has to be based on the diagnosis and not serology. Ideally everyone would have a biopsy and that would be used as the gold standard. Though the sensitivity of biopsy is highly dependent upon the site and number of biopsies taken (but data are relatively sparce). NICE did not define "weakly positive". Presumably because, as with many autoantibody tests, there is poor between assay comparability, standardisation and linearity. Having both TTG and EMA positive improves the PPV and helps make the case for biopsy but is still not diagnostic. In patients with significant signs or symptoms of coeliac, TTG+/EMA- patients should still be considered for biopsy. Or at least be reviewed and retested in 6-12m. As a result in many cases EMA may be a distraction rather than a help. However, I think it's useful in patients with a low pre-test probability and weak pos TTG. EMA usefulness will depend upon the specificity of the TTG test in use. That difficult balance between FP and FN results! Local data is great for verifying manufacturer cut-offs and, like you, we've done this for TTG. Sample size is an issue if you want to use it to set cut-offs from scratch though because confidence intervals tend to be quite wide. This takes many years of ongoing clinical audit and case finding/follow up. We work back from an annual trawl of positive biopsy samples but with only about 70% having a biopsy and a fall-off in the proportion of children having biopsy, there is a risk of significant selection bias. If I was starting from scratch and didn't have a verified reference cut-off for TTG, I'd aim for a 99th percentile too (easier said than done!) as a starting point. Immunofluorescence tests like EMA require local optimisation but I suspect many centres think this is the responsibility of the manufacturer. Like you, we find that raised IgA for whatever reason (even big paraproteins) only occasionally gives false (always weak) pos TTG. For this reason we routinely check IgA level if TTG is weak pos and AMA is negative. Thanks for raising some interesting issues! Best Wishes Steve Prof Stephen Holding PhD FRCPath Consultant Clinical Scientist (Clinical Immunology) Honorary Professor, Hull York Medical School Blood Sciences, Pathology Dept, Hull and East Yorkshire Hospitals, Anlaby Road, Hull, HU3 2JZ • +44 (0)1482 607710 -----Original Message----- From: Richmond Katherine (RLZ) [mailto:[log in to unmask]] Sent: 18 June 2018 11:09 To: 'Steve Holding' Subject: RE: Coeliac serology Hi- I would reply publicly but cant find my password! This is something that I feel very strongly about.... Cut-offs should be chosen to match biopsy results , I do not consider the EMA test to be the gold standard I'm afraid. I reluctantly re-introduced endomysial antibody testing to support the paediatric guidelines as regards diagnosis and avoiding biopsy. If asked I would be more inclined to say we should be using a 99th centile approach for TTG. We introduced the Phadia Varelisa assay years ago and we used to send positive TTG for endomysial testing. We audited the results very closely. We took a years worth of negative and positive biopsy data and matched it to serology in order to try and minimise selection bias We found a number of things. 1. Of a goup of patients with borderline TTG results (4-8) who proved on biopsy to have coeliac- 50% were negative on endomysial testing. The agreement with the endomysial result from the referral lab was not ideal and could have led to misdiagnosis. We found that over the next few years the agreement improved ... implying that the endomysial assays were subject to variation- and possibly that cycles of TTG-endomysial-clinical audit were highlighting deficiencies.... 2. We found that the cut-offs provided by the manufacturer were too high- probably based on selection bias using samples and patients selected on the basis of endomysial testing results. We now use cut-offs of approximately half that recommended by Phadia. Phadia recommend that normal is up to 4. We use 2-4 as borderline. As part of the move to the Immunocap we re-validated these cut-offs, and found that 6/7 children biopsied with levels between 2 and 4 were found to have coeliac disease. 3. In children- results over 1 are mostly observed in type 1 diabetics, children with strong family history of coeliac , or treated coeliac. In children with type 1 diabetes having annual screens you can see the TTG levels gradually rising over the years. 4. We do not find patients with liver disease to be a major problem. The ones that I have seen have very marginally elevated TTG associated with very high total IgA levels. I don't see endomysial antibody testing as adding value since I believe they would be negative even for a true coeliac at this kind of level. Regards Katherine Richmond Lead Principal Biochemist Shrewsbury and Telford Hospital NHS Trust Mytton Oak Rd Shrewsbury SY3 8XQ 01743 261157 -----Original Message----- From: Clinical biochemistry discussion list [mailto:[log in to unmask]] On Behalf Of Steve Holding Sent: 17 June 2018 18:50 To: [log in to unmask] Subject: Re: Coeliac serology We check all positive TTG with EMA. About 2% of our requests are positive so this amounts to a slide a week. This would be no different if we just confirmed 'weakly positive' results. Difficult to define 'weakly positive' but I use <30 u/mL (double the lower limit of the ref. interval, which is <15 u/mL). We use Biorad Bioplex 2200 TTG-IgA/G assays. Best Wishes Steve Steve Holding PhD FRCPath Consultant Clinical Scientist (Clinical Immunology) Honorary Professor, Hull York Medical School Blood Sciences, Pathology Dept, Hull and East Yorkshire Hospitals, Anlaby Road, Hull, HU3 2JZ o +44 (0)1482 607710 -----Original Message----- From: Clinical biochemistry discussion list [mailto:[log in to unmask]] On Behalf Of CLIFFORD-MOBLEY, Oliver (UNIVERSITY HOSPITALS BRISTOL NHS FOUNDATION TRUST) Sent: 15 June 2018 17:21 To: [log in to unmask] Subject: Coeliac serology Dear mailbase, NICE NG20 recommends confirming weakly positive IgA TTG antibodies with IgA endomysial antibodies (EMA). How have other centres defined and verified the TTG weakly positive range? We recently switched TTG assay and during our verification found that some TTG results above the manufacturer's stated weakly positive range were negative by EMA. Therefore we are currently confirming all positive TTG by EMA and not defining a 'weakly positive' range - are any other centres doing the same thing? The alternative we could consider is to define the upper limit of the TTG weakly positive range such that it has 100% concordance with EMA positivity. However the question that arises from this is whether EMA should be considered the gold standard: I understand that it has a greater specificity than TTG for CD but I also believe that a false negative EMA is still possible. Many thanks, Oliver Clifford-Mobley FRCPath Principal Clinical Scientist Department of Clinical Biochemistry University Hospitals Bristol NHS Foundation Trust T: 0117 342 7833 ******************************************************************************************************************** This message may contain confidential information. If you are not the intended recipient please inform the sender that you have received the message in error before deleting it. 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Please note, archived messages are public and can be viewed via the internet. Views expressed are those of the individual and they are responsible for all message content. ACB Web Site http://www.acb.org.uk Green Laboratories Work http://www.laboratorymedicine.nhs.uk List Archives http://www.jiscmail.ac.uk/lists/ACB-CLIN-CHEM-GEN.html List Instructions (How to leave etc.) http://www.jiscmail.ac.uk/