Hyaluronan (HA) is widely detected in biological samples and its concentration is most commonly determined by the use of a labeled specific HA binding protein (aggrecan G1-IGD-G2, HABP), employing membrane blotting and sandwich enzyme-linked immunosorbent assay (ELISA)-like methods. However, the detected signal intensity or the quantified value obtained by using these surface-based methods is related to the molecular mass (M) of HA, especially for HA in the low M range below ~150 kDa. At the same mass or mass concentration, higher M HA gives a higher signal than lower M HA. We have experimentally determined the quantitative relationship between the M of HA (in the range 20-150 kDa) and the relative signal intensity in comparison with a standard HA, in a sandwich ELISA-like assay. An M-dependent signal correction factor (SCF) was calculated and used to correct the signal intensity, so that the corrected concentration value would more accurately reflect the true HA concentration in solution. The SCF for polydisperse low M HA was also calculated and compared with experimental results. When the molecular mass distribution of an HA sample is determined by a method such as gel electrophoresis, then its appropriately averaged SCF can be calculated and used to correct the signal in sandwich ELISA to obtain a more accurate concentration estimation. The correction method works for HA with M between ~150 and 20 kDa, but lower M HA is too poorly detected for useful analysis. The physical basis of the M-dependent detection is proposed to be the increase in detector-accessible fraction of each surface-bound molecule as M increases.

Background Most of hematology laboratories in Saudi Arabia utilize the reference intervals (RIs) provided by instrument manufacturers. This study aimed to define RIs of hematological parameters for adult population in the western region of Saudi Arabia and to explore their specific features from an international perspective. Method This study was conducted according to the harmonized protocol of IFCC Committee on RIs and Decision Limits. Blood samples collected from 409 healthy Saudi males and females adults were analyzed for complete blood count (CBC) by using Cell-Dyn Sapphire analyzer and for iron profile by using Architect analyzers. The needs for RIs partitioned by sex and age was based on standard deviation ratio (SDR) and/or bias ratio (BR). RIs were derived parametrically with/without application of the latent abnormal values exclusion method (LAVE). Results Based on thresholds of SDR≥0.4 and/or BR≥0.57, RIs were partitioned by sex for red-blood cell count, hemoglobin, hematocrit, red cell distribution width, erythrocyte sedimentation rate, iron, transferrin, ferritin, eosinophil, platelet, plateletcrit, etc. Partitioning by age was not necessary for any of the analytes. LAVE procedure caused appreciable changes in RI limits for most erythrocyte and iron parameters but not for leukocyte parameters. Comparable to other non-IFCC studies on CBC RIs, the RBC and hematocrit (Ht) ranges have shifted to a higher side in both genders. After applying the LAVE method, the male and female RIs for Hb were 4.56 to 6.22 ×106/μL and 3.94 to 5.25 ×106/μL respectively while RIs for Ht were 40.2 to 52.0% and 33.6 to 44.5% respectively. Conclusion LAVE method contributed to reducing the influence of latent anemia in deriving RIs for erythrocyte related parameters. Using the up-to-date methods, the RIs of CBC determined specifically for Saudis will help to improve the interpretation of test results in medical decision making.