Modification and validation of the test procedure for determination of sub-visible particulate matter in parenteral solutions, using the Coulter method

The State Pharmacopoeia of the Russian Federation, 14th edition provides for determination of sub-visible particles (less than 100 µm in size) in parenteral dosage forms using the Coulter method, in addition to the light obscuration particle count test and microscopy. However, the proposed 100 µm aperture tube does not enable assessment of the whole range of sub-visible particle sizes. Therefore, research is needed to find optimal test conditions for determination of sub-visible particulate matter by the Coulter method. The aim of the study: modification of the Coulter-based procedure using a 200 µm aperture tube, and performance of validation studies. Materials and methods: Multisizer 4e Coulter counter, suspensions of reference latex particles (10 µm, 20 µm, and 43 µm), and a particulate count reference standard containing 0.998 × 106 particles/mL were used in the study. The following parameters were assessed during validation: accuracy, repeatability, linearity. Results: the study confirmed the feasibility of using the modified Coulter-based procedure with a 200 µm aperture tube. The following values were obtained during validation of the modified test procedure: accuracy was 5.3% (deviation from the mean value) as compared to the particulate count reference standard, and 4.2% as compared to the light obscuration method. Repeatability was 1% (relative standard deviation) for the particle concentration of approximately 10000 per 1 mL, and 7.6% for the particle concentration of approximately 300 per 1 mL. The study demonstrated the linearity of the procedure, the linear correlation coefficient was more than 0.99. Conclusions: the studied validation parameters of the modified test procedure were shown to comply with the acceptance criteria. The modified test procedure will enable assessment of the whole range of sub-visible particle sizes when testing parenteral solutions for particulate contamination: sub-visible particles.

Observational Study on Variation of Longitudinal Platelet Counts in Calves over the First 14 Days of Life and Reference Intervals from Cross-Sectional Platelet and Leukocyte Counts in Dairy Calves up to Two Months of Age

Platelet and leukocyte count reference intervals (RIs) for cattle differ by age and while adult RIs are known, RIs for calves are studied less. The aims of this observational study are to evaluate variation of platelet counts of Holstein Friesian calves over the first 14 days of life and to propose RIs for platelet and leukocyte counts of Holstein Friesian calves aged 0–60 days. In a longitudinal study, 19 calves were blood sampled 17 times, in the first 14 days of their lives. Blood was collected in a citrate blood tube and platelet counts were determined. We assessed the course of platelet counts. In a field study, 457 healthy calves were blood sampled once. Blood was collected in an EDTA blood tube and platelet and leukocyte counts were determined. The RIs were calculated by the 2.5 and 97.5 percentiles. Platelet counts started to increase 24 h after birth (mean platelet count 381 × 109/L ± 138 × 109/L) and stabilized after five days (mean platelet count 642 × 109/L ± 265 × 109/L). In calves up to six days of age, platelet counts were lower than in calves older than five days. In conclusion, the RIs of platelet and leukocyte counts in calves were wider in range than the RIs for adult cattle, therefore, calf specific RIs for platelet and leukocyte counts should be used. From 6 until 60 days of age, we propose an RI for platelet counts of 287–1372 × 109/L and for the first 60 days of life an RI for leukocyte counts of 4.0–18.9 × 109/L.