CLEANING VALIDATION OF TWO TYPES OF EQUIPMENT
Considering the results shown above, the TOC test was used to assess the cleaning of bioproduction equipment used in two types
of manufacturing processes. The first set of equipment was used to obtain a recombinant protein from E. coli bacterial cells as host, and in the second scenario, S. cerevisiae yeast cells are used to express a heterologous protein. In both manufacturing systems, one fermentation vessel and one chromatography
column were taken as examples.
Before the cleaning process was performed, the procedures used (who is responsible for performing and approving them, acceptance
criteria, revalidation) were documented in detail.1 Then, the cleaning process was performed, as described in preapproved protocols.11–13
The results of three independent cleaning processes for both manufacturing systems are shown in Tables 3 and 4. For the fermenter
vessel, cleaning validation results from rinse samples showed a reduction factor of three orders of magnitude, with final
values ranging from 1.27 x 103 to 2.25 x 103. The values from the harvest port swabbing samples showed the same trend as for rinse samples, with a three orders of magnitude,
and final values ranging from 1 x 103 to 3.73 x 103 .
Table 3. Cleaning validation results (TOC, in ppb) for fermenters used in E. coli and S. cerevisiæ production processes
On the other hand, the results of chromatography column cleaning showed, in general, lower reduction factor values compared
with those for fermenter cleaning. Those results were expected, knowing that the chromatography system is in contact with
cleaner material than that seen in fermentation steps.
The final samples (rinse and swabbing samples) of both manufacturing systems showed TOC values as low as 22 ppb (close to
the LOQ, Table 1). The TOC value obtained for the water used to clean the biomanufacturing system was used as a reference
The results of the swabbing recovery study (Table 2) showed that the levels of organic charge reduction after cleaning reported
in this work, showed a high level of cleaning efficiency. This proposed cleaning validation strategy can be applied to both
single and multiproduct manufacturing facilities.
Table 4. Cleaning validation results (TOC, in ppb) for chromatography columns used in E. coli and S. cerevisiæ production
A general strategy for assessing cleaning validation using the total organic carbon (TOC) test was applied. The qualification
of the TOC measurement system showed a precision of 1.27%, 1.3% of accuracy, and a detection limit of 6.30 ppb of TOC.
A linear correlation between cells and TOC values was demonstrated with this analytical system. This study showed that in
both 27 ppb and 16 ppb of TOC, ~106
E. coli cells and 103
S. cerevisiae cells were present, respectively. The swab recovery assessment showed recovery factor values ranging from 90 to 110% using
stainless steel and borosilicate surfaces and SK, EGF, IFNα2bHu-r, as well as both E. coli cells and S. cerevisiae as biological matrices, and purified water as a recovery solvent. It also showed that applying the polyethylene glycol conjugation
process to IFNα2bHu-r does not affect its interaction with borosilicate surfaces. Given the cleaning reduction factors values,
the final TOC values for the studied systems, the agreement between them, and the TOC values from final samples and for the
water used to clean each system, the strategy described here can be a useful tool in similar cleanliness validation strategies.
The authors want to thank Magaly García Blanco for proofreading this manuscript and the epidermal growth factor and interferon
manufacturing personnel from the CIGB for supplying the cleaning validation samples used in this work.
Julio César Sánchez is the head, and Raudel Sosa and Meily Sánchez are technicians in the process control department, Rebeca Bouyon Albarran is a research scientist in the epidermal growth factor department, Luciano Hernández and Marbel Ramos Alfonso are research scientists in the interferon manufacturing department, Alexis Musacchio Lasa is a research scientist in the bioinformatics department, and Leopoldo Núñez is a technical service assistant, all at the Center for Genetic Engineering and Biotechnology, Havana, Cuba, +537.271.6022
(ext. 7101), firstname.lastname@example.org