P.S., M.H., P.M. the suitability of plastic microtubes for filling small quantities. The stability and quality of the dried material was assessed after an accelerated degradation study using a toxin neutralization test and size exclusion HPLC. While microtubes have shown some promise in the past for use in the lyophilization of some biological materials, issues with stability may arise when more labile materials are freeze-dried. We demonstrate here the microtube format is definitely unsuitable for ensuring the stability of this monoclonal antibody. sodium dodecyl sulfate in 50% dimethylformamide, pH 4.7 (100 L/well). The plates were returned to the incubator over night to allow for the complete extraction and solubilization of the coloured product, and the OD was read at 570 nm (Vmax, Molecular Products, Wokingham, UK). The end-point for each mAb sample was identified as the last dilution showing neutralization of the toxin (defined as OD 50% of the cell only control wells). 3. Results and Discussion 3.1. Results of the Freeze Drying For the SS-701 (diagnostic formulation), Number 1 shows the freeze-drying cycle used. Ozagrel(OKY-046) This was a two-day cycle with freeze at ?50 C, main drying at ?35 C for 20 h, followed by a ramp over 8 h to secondary drying at 30 C for 7 h using the pilot level Telstar Lyobeta freeze dryer. Open in a separate window Number 1 Freeze drying method and profile for SS-701 diagnostic formulation in Telstar Lyobeta freeze dryer. The run profile is also demonstrated in Number 1. The programmed cycle was completed as expected, suggesting that the product dried properly over 2 days. A convergence of the vacuum pirani and baratron gauges midway through the primary drying step indicated the end of the primary drying, as the pirani gauge is sensitive to water vapor coming off during main drying. The restorative formulation was freeze dried over two days in the Virtis Genesis freeze dryer. This is a pilot level freeze dryer similar in specification to the Telstar Lyobeta freeze dryer utilized for the diagnostic formulation. The freeze-drying cycle was identical, and the cycle was completed as programmed. 3.2. Freeze Dried Appearance Diagnostic versus Restorative Formulation The appearance of both the diagnostic and the restorative formulations was assessed after freeze drying in terms of the robustness of the freeze-dried cake formed. Physique 2a,b show the freeze-dried products of the diagnostic and the therapeutic formulation in both ampoules and microtubes. The therapeutic formulation produced a more robust freeze-dried cake, whereas the diagnostic formulation produced a shrunken, crumbly cake, which began to break up on handling. Open in a separate window Physique 2 (a) Diagnostic Formulation freeze dried appearance in ampoules and microtubes. (b) Therapeutic Formulation freeze-dried appearance in ampoules and microtubes. 3.3. SE-HPLC Results The SE-HPLC data are presented in Table 1 as % monomer relative to the ?20C samples at 1 and 16 week timepoints. In the microtube format, a decrease in the monomer peak was observed at elevated storage temperatures from about 94% down to 45% of the ?20 C value. At the highest storage temperature, the pre-monomer peaks increased in size and several other peaks also occurred after 20 min, suggesting the product is unstable in microtubes. On the other hand, the ampoules showed good stability after 16 weeks storage for diagnostic and therapeutic formulations (Table 1) when compared to the microtubes. This is exhibited by the fact that this % monomer content was no less than 94% of the Ozagrel(OKY-046) ?20 C value for all the storage temperatures for the product filled in the ampoules. However, the microtubes showed a high Mouse monoclonal to CD25.4A776 reacts with CD25 antigen, a chain of low-affinity interleukin-2 receptor ( IL-2Ra ), which is expressed on activated cells including T, B, NK cells and monocytes. The antigen also prsent on subset of thymocytes, HTLV-1 transformed T cell lines, EBV transformed B cells, myeloid precursors and oligodendrocytes. The high affinity IL-2 receptor is formed by the noncovalent association of of a ( 55 kDa, CD25 ), b ( 75 kDa, CD122 ), and g subunit ( 70 kDa, CD132 ). The interaction of IL-2 with IL-2R induces the activation and proliferation of T, B, NK cells and macrophages. CD4+/CD25+ cells might directly regulate the function of responsive T cells level of monomer loss down to about 30% of the total integrated area for both the diagnostic and the therapeutic preparations after 16 weeks when stored at 45 C. The freeze-dried mAb was also tested after 1 week against the liquid material and no degradation was observed for any of the container formats immediately after freeze drying. Table 1 Diphtheria antitoxin % monomer content of freeze-dried DATMAB after storage at elevated temperature for 1 and 16 weeks. Data are the % monomer relative to ?20C samples. thead th align=”center” valign=”middle” style=”border-top:solid thin;border-bottom:solid thin” rowspan=”1″ colspan=”1″ Diagnostic FormulationTubes /th th align=”center” Ozagrel(OKY-046) valign=”middle” style=”border-top:solid thin;border-bottom:solid thin” rowspan=”1″ colspan=”1″ 1 Week /th th align=”center” valign=”middle” style=”border-top:solid thin;border-bottom:solid thin” rowspan=”1″ colspan=”1″ 16 Weeks /th /thead +20 C96.3477.33+37 C97.8476.46+45 C89.5044.36 Diagnostic FormulationAmpoules 1 Week 16 Weeks +20 C100.7698.81+37 C101.3998.03+45 C100.9394.88 Therapeutic FormulationTubes 1 Week 16 Weeks +20 C99.7594.96+37 C99.0276.34+45 C92.4048.41 Therapeutic FormulationAmpoules 1 Week 16 Weeks Ozagrel(OKY-046) +20 C100.30102.07+37 C100.17102.16+45 C99.37102.26 Open in a separate window.
