recent years in hospital laboratories and blood banks. We describe a simple and rapid ... Sant Joan de Reus, Carrer de Sant Joan s/n, 43201 Reus, Spain.
CLIN. CHEM. 40/4, 625-629 (1994)
Enzymes and Protein Markers
#{149}
Automated Latex Agglutination Immunoassay of Serum Ferritin with a Centrifugal Analyzer Josep Maria Sim#{243},’ Jorge Joven, Xavier Clivill#{233}, and Teresa Sans A considerable demand for convenient, rapid, inexpensive assays of ferritin in serum has been generated in recent years in hospital laboratories and blood banks. We
metric immunoassays may be extended by using light-scattering properties of the immunoaggregates tained by attaching the antibody to latex particles,
describe a simple and rapid particle-enhanced turbidimetnc immunoassay suitable for routine application in a Monarch 2000 centrifugal analyzer with commercially available reagents. This fully automated assay (y) requires no pretreatment of sample, and correlation with a two-step sandwich ELISA (x) is excellent (y = 1.01 8x + 0.397, S = 0.027). The analytical range extends from 5 to 900 pgIL. lntraassay imprecision (CV) ranged from 1.1% to 5% for various specimen concentrations. Interassay imprecision ranged from 2.2% for above-normal concentrations (755 g/L) to 9.5% for low concentrations (39 p.gIL). No specimen-related carryover was detected. The method has been useful in our predeposit autologous blood transfusion program for rapid assessment of iron status in patients undergoing repeated phlebotomies.
so-called microparticle-enhanced turbidimetric immunoassay or latex agglutination immunoassay (12, 13). Therefore, serum ferritin as well as other analytes may be reliably measured by this method, with advantages of practicality, decreased operating time, and cost in
Indexing Terms: pared/pro teins
iron/turbidimetric
immunoassay/ELISA
com-
Ferritin, the intracellular iron-storage protein, is also found in serum or plasma (1); its determination is considered to be the most reliable method for evaluating iron stores (2, 3). Measurements reflecting iron stores are the most sensitive indices of iron status because iron stores are depleted during the evolution of iron deficiency, before iron in serum and erythrocytes is significantly affected (4). Increased awareness of the biochemical function of ferritin, the clinical consequences of iron depletion, and the endorsement of autologous blood transfusion (5) with repeated phiebotomies have generated considerable demand for a convenient, routine, rapid, and inexpensive assay of ferritin in serum. Several methods for measuring ferritin have been reported, including immunoradiometric assays (IRMAs) (1), RIAs (6), ELISAs (7), fluorescence polarization immunoassays (8), and chemiluminescence assays (9, 10). Hazards associated with radiochemicals have reduced the use of both IRMA and RIA, but nonisotopic methods do not represent an improvement in time requirements or in the need for special, often dedicated, instruments. Conversely, turbidimetric immunoassay is well established as a homogeneous assay for the rapid quantification of analytes at concentrations not suitable for ferritin measurements (milligrams per liter or more) (11). However, the concentration range applicable to turbidiCentre de Recerca Biom#{232}dica, Laboratoris Clinics, Hospital de Sant Joan de Reus, Carrer de Sant Joan s/n, 43201 Reus, Spain. for correspondence. Fax 34-77-323690. Received May 11, 1993; accepted December 8, 1993.
the obthe
comparison with other nonisotopic methods. Here we describe an assay adapted for use in a centrifugal analyzer and compare it with a well-established ELISA.
Materials and Methods Apparatus We used a Monarch 2000#{174} Chemistry System, a centrifugal analyzer manufactured by Instrumentation Laboratory (Milan, Italy). For comparison studies we used the Enzymum Test System ES-300#{174}, a modular batch-analyzer system for the automatic performance of tests with reagents from Boehringer Mannheim (Marburg, Germany). Sample and Control Specimens We used Moni-Trol#{174} X Chemistry Control Level I/lI (cat. no. 160573; Baxter Dade, Basel, Switzerland) and different pools of serum to study the analytical variables. For comparison studies, we obtained venous blood from 91 volunteers (47 men, 44 women, ages 25-50 years), including normal subjects and patients with known disorders of iron metabolism. To assess possible interferences of hypergammaglobulinemia in the procedure, we also performed a comparison with specimens from 23 patients with abnormally high y-band content, as identified by agarose gel electrophoresis (15 patients with liver cirrhosis, 6 patients with arthritis, and 2 patients with polyclonal gammopathy). Blood was allowed to clot at 37#{176}C, and serum was obtained by centrifugation at 700g for 10 mm and stored at -80#{176}C until assay. All procedures were done in accordance with the ethical standards of our institution. Reagents Latex microparticles sensitized to ferritin and standards. Al gIL suspension in ammoniacal buffer of polystyrene particles (diameter 0.1 jim, density 1.040 kgfL) coated with covalently attached F(ab’)2 fragments of rabbit polyclonal IgG anti-human ferritin (30-300 ng/ 2) was obtained from Eiken Chemical Co., Tokyo, Japan (LA ferritin, cat. no. 8X2VLC14). The latex microparticles had a refractive index of 1.59, and the mean absorbance of the reagent at 600 nm was 1069 mA, with minor variations between lots (
