Thesis (MMedSc)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: Background: Serum free light chains (FLC) are associated with imbalances in heavy and light
chain production. Abnormal FLC ratios have been associated with risk of progression in certain
diseases. Automated assays are available for their determination and they are used in the followup
and management of patients with monoclonal gammopathies. Acceptable imprecision,
specificity, accuracy and reproducibility between reagent batches is required to prevent under- or
overestimation. Method validation is a standard process in every good laboratory to judge the
acceptability of a new method. Reference intervals have been established in an older population,
but it was considered important to verify these in our population. HIV is associated with B-cell
dysfunction. As B-cell abnormalities are associated with disorders leading to monoclonal
gammopathies, we postulated that the FLC levels and FLC ratio would be abnormal in HIV
infected individuals.
Methods and materials: Controls and pooled patient samples were used for the method validation
study which included imprecision studies, linearity, recovery and interference studies, and
method comparison studies, the latter compared our method to the same method used in another
laboratory. For the reference interval study, blood was obtained from 120 healthy subjects. The
following blood tests were performed: total protein, IgG, IgA, IgM, creatinine, protein
electrophoresis, kappa FLC and lambda FLC. Using the kappa and lambda FLC results, a FLC
ratio was determined. Three hundred and sixty-nine HIV positive subjects were then studied. The
same tests were performed, as well as CD4+ counts and viral loads on the majority of them.
Results: For the method validation study, precision, linearity and recovery was acceptable.
Minimal interference was observed with haemolysis, lipaemia, bilirubin and rheumatoid factor.
Our method showed comparable performance with the established method. For the reference
interval study, all the creatinine values were normal, as were serum protein values. The serum
protein electrophoreses were independently reviewed by 3 pathologists. Most were normal, with
a few polyclonal increases seen, but no definite monoclonal bands. The 95% reference intervals
for FLC’s as well as the FLC ratio were not statistically significantly different to the
manufacturer’s recommendations. When examining the HIV positive study population, we found that FLC and FLC ratio were influenced by markers of HIV disease severity, such as CD4+
count, IgG, viral load, use of antiretroviral treatment and abnormal serum protein
electrophoreses.
Conclusion: The validation study of FLC showed excellent precision, acceptable bias, good
linearity, good recovery and minimal interference, allowing routine introduction of the test. The
95% reference intervals obtained for our population were slightly higher than those
recommended by the manufacturer. However, as most of the values fell within the
manufacturer’s limits, we could accept the manufacturer’s recommended cut-offs. We found that
FLC levels were definitely influenced by markers of HIV disease severity in our population and
we postulate that they may be of use for follow-up of patients with HIV. / AFRIKAANSE OPSOMMING: Agtergrond: Serum vry ligte kettings (VLK) word geassosieer met ‘n wanbalans van ligte en
swaar ketting produksie. Abnormale VLK ratios is geassosieer met ‘n risiko van verloop in
sekere siektes. Geoutomatiseerde laboratorium toetse vir VLK is beskikbaar vir hul bepaling en
word gebruik om pasiënte met monoklonale gammopatieë op te volg en te behandel.
Aanvaarbare impresisie, spesifisiteit, akkuraatheid en herhaalbaarheid tussen reagens besendings
is belangrik om onder- of oorbepaling te verhoed. Metode validasie is ’n standaard proses in elke
goeie laboratorium om die aanvaarbaarheid van ’n nuwe metode te bepaal. Verwysingswaardes
is al bepaal in ’n ouer populasie. Ons het besluit om die verwysingswaardes in ons populasie te
bepaal. Mens-immuungebrekvirus (MIV) word geassosieer met B-sel disfunksie. Omdat B-sel
abnormaliteite geassosieer word met afwykings wat tot monoklonale gammopatieë lei, het ons
gepostuleer dat die VLK vlakke en VLK ratio abnormaal sal wees in MIV geïnfekteerde persone.
Metodes en Materiale: Kontroles en pasiënt monsters is gebruik vir die metode validasie studie
wat impresisie studies, lineariteit, herwinning, inmenging en metode korrelasie studies ingesluit
het. In laasgenoemde geval is ons metode met dieselfde metode van ’n ander laboratorium
vergelyk. Vir die verwysingswaardes studie is 120 gesonde persone se bloed gebruik. Die
volgende toetse is bepaal: totale proteïen, IgG, IgA, IgM, kreatinien, proteïen elektroferese,
kappa en lambda VLK. Die VLK ratio is bepaal deur die kappa en lambda resultate te gebruik.
Driehonderd nege en sestig MIV-positiewe pasiente is gebruik vir die studie. Dieselfde toetse
was gedoen, asook CD4+ tellings en virale ladings op die meerderheid van pasiente.
Resultate: Vir die metode validasie studie, was presisie, lineariteit en herwinning aanvaarbaar.
Minimale inmenging van hemolise, lipemie, bilirubien en rumatoïede factor is waargeneem. Ons
metode het goed gekorreleer met die bepaalde metode. Die serum kreatinien en serum totale
proteïen waardes was normaal tydens die verwysingswaardes studie. Die serum proteïen
elektroferese was onafhanklik beoordeel deur 3 patoloë. Die meeste was normaal met enkele
poliklonale verhogings, maar geen definitiewe monoklonale bande nie. Die 95% verwysings
intervalle vir VLK en VLK ratio het nie statisties betekenisvol verskil van die vervaardiger se
aanbevelings nie. In die studie van die MIV-positiewe studie populasie, het ons gevind dat VLK en VLK ratio beïnvloed word deur merkers van ernstige MIV siekte, soos CD4+ telling, IgG,
virale lading, die gebruik van antiretrovale medikasie en abnormale serum proteïen elektroferese.
Gevolgtrekking: Die validasie studie van VLK het uitstekende presisie, aanvaarbare
partydigheid, goeie lineariteit, goeie herwinning en minimale inmenging gewys, wat die roetine
instelling van die toets toegelaat het. Die 95% verwysingsintervalle wat vir ons populasie bepaal
is, was effens hoër as die vervaardiger se aanbeveling. Die meeste van die waardes het egter
binne die vervaardiger se limiete geval, dus kon ons die vervaardiger se afsnypunte aanvaar. Ons
het gevind dat VLK vlakke definitief beïnvloed word deur merkers van die ernstigheidsgraad van
MIV siekte in ons populasie en ons postuleer dat VLK van waarde kan wees met die opvolg van
MIV pasiente. / NHLS / Harry Crossley for funding obtained
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:sun/oai:scholar.sun.ac.za:10019.1/20173 |
| Date | 03 1900 |
| Creators | Germishuys, Jurie J. |
| Contributors | Zemlin, A. E., Erasmus, R. T., Stellenbosch University. Faculty of Health Sciences. Dept. of Pathology. Chemical Pathology. |
| Publisher | Stellenbosch : Stellenbosch University |
| Source Sets | South African National ETD Portal |
| Detected Language | Unknown |
| Type | Thesis |
| Format | 101 p. : col. ill. |
| Rights | Stellenbosch University |
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