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Sean,
The fact that you are seeing this problem
with plasma ( but not serum ) makes me wonder if some of these plasma samples contain platelets, which could be
lysed during the assay, releasing LD.
Do you remove all platelets from plasma
with your current combination of g force and centrifugation time ( check platelet
count of plasma samples using haematology analyser )?
The patient with idiopathic
thrombocytopoenic purpura, however, is difficult to explain.
How hard would it be omit NADH from your
LD assay mixture, and monitor absorbance changes for serum and plasma samples
causing you problems?
I am sure that Beckman could provide you
with the reagent formulation in confidence.
This would then allow you to resolve whether
the interference was chemical or optical.
We had a similar problem many years ago.
I have attached the paper summarising our
investigations.
Hope this helps, and it would be
interesting to know what you find.
Kind regards,
Michael Peake,
Flinders Medical Centre,
From: Clinical
biochemistry discussion list [mailto:[log in to unmask]] On Behalf Of Sean Maguire
Sent: Thursday, 9 July 2009 11:12
PM
To:
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Subject: LDH (pyruvate to lactate)
nonlinear reactions
Dear colleagues,
Here in the Biochemistry lab of the
Absorbances of the reaction can be viewed after the assay is
over by highlighting LDH and then ABS.
We get the occasional plasma where the absorbance in the
reading time is not linear.
The reaction , instead of being linear in the read time
curls upwards at the end as if NADH was being produced from another source.
We dilute with a control (1:1) and then we get linearity and
also higher answers for the patient sample which we then report.
Serum samples on these same patients with the the
interference are linear in the read time.
The interference in plasma seems to disappear after about 5
hours which makes it hard to examine.
The patients in whom we see this interference are invariably
patients with cancer or on cancer treatment (most from Haematolgy OPD).
One patient had idiopathic thrombocytopoenic purpura and was
apparently on no unusual treatment.
We could switch to serum for all LDHs as a solution.
Can I get your advice as to where the interference is coming
from? No one drug is implicated in all the patients...
Many thanks.
Dr. Sean Maguire,
Principal Biochemist,