Original Research Cleaning validation of different cleaning procedures for monoclonal antibodies on stainless steel Jana Thissen, MSc a,b , Lars M. H. Reinders a,c , Jochen Tuerk a,d,e , Martin D. Klassen, MSc a, * Abstract Introduction: Despite their growing importance in tumor treatment, monoclonal antibodies (mAbs) are still underrepresented in investigations on the efficiency of cleaning procedures. Effective cleaning routines are essential for pharmacies to minimize occu - pational exposure as well as proteinaceous surface contamination which otherwise can result in microbial growth. This study focused on the efficiency of different cleaning agents (alkaline, alcohol) and procedures (single pass, multipass) to remove various mAbs from stainless steel. Methods: Stainless steel surfaces were contaminated with known concentrations of various mAbs. Specifically, this study involved the use of bevacizumab, cetuximab, daratumumab, omalizumab, rituximab, and trastuzumab. Then, different standardized cleaning procedures were applied. This was followed by wipe sampling, enzymatic digestion of the wipe extracts, and analysis of the digested wipe extracts by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) to investigate the cleaning efficiency of the different cleaning procedures. Results and Discussion: The cleaning performance of the alkaline cleaning solutions investigated was significantly higher than of 70% isopropyl alcohol (70% IPA) in the removal of various mAbs from stainless steel. Isopropyl alcohol has a low cleaning efficiency for the removal of mAbs on stainless steel. The cleaning procedures evaluated in this study are contextualized within a cleaning protocol recommended by the European Society of Oncology Pharmacy and the contamination control strategy outlined in EU GMP Annex 1. Conclusion and Outlook: In this study, the cleaning efficiency of different solutions and procedures for removing monoclonal antibodies from stainless steel was evaluated for the first time to our knowledge. Wipe sampling and analysis of monoclonal antibodies on stainless steel surfaces could be successfully implemented. It was demonstrated that multipass cleaning three-cycle two-agent procedure with an alkaline cleaner followed by alcohol cleaner results in complete removal of the applied monoclonal antibodies on stainless steel. Future studies could focus on assessing further cleaning agents and substrates such as glass. Keywords: cleaning efficiency evaluation, occupational safety, monoclonal antibodies, stainless steel 1. Introduction In tumor treatment, monoclonal antibodies (mAbs) gained in - creasing importance over the past 20 years. The use of personalized medicine is widespread in cancer therapy, which requires trained pharmaceutical staff to dose patient-specific preparations under aseptic conditions. Although efficient cleaning routines for cytostatic agents have been well studied, [1 – 6] there is a lack of data on cleaning strategies for monoclonal antibodies. However, according to the contamination control strategy (CCS) outlined in EU GMP Annex 1, [7] cleaning procedures must be evaluated. In addition, efficient cleaning strategies are crucial when handling monoclonal antibodies to prevent occupational exposure. The This research was funded by Sch ¨ ulke & Mayr GmbH. Special thanks to Andr ´ e Lembke and Christin Becker for their valuable contributions to the study design. Sch ¨ ulke & Mayr GmbH provided perform sterile Mucasol, perform sterile alcohol IPA and perform sterile mix dry wipes used in this study free of charge. The company was not involved in data collection, statistical analysis, interpretation of results, manuscript preparation. The authors affirm that the study was conducted impartiall y and in accordance with the highest scientific standards. All authors declare no other financial or personal conflicts of interest. During the preparation of this work the authors used ChatGPT (Version GPT-4, OpenAI, San Francisco, USA) in order to improve the readability and clarity of the text. After using this tool/service, the authors reviewed and edite d the content as needed and take full responsibility for the content of the published article. a Institut f ¨ ur Umwelt & Energie, Technik & Analytik e.V. (IUTA), Duisburg, Germany, b Heinrich Heine University D ¨ usseldorf, Faculty of Mathematics and Natural Sciences, Institute of Pharmaceutics and Biopharmaceutics, D ¨ usseldorf, Germany, c Romaco Innojet GmbH, Steinen, Germany, d Centre for Water and Environmental Research (ZWU), University Duisburg-Essen, Essen, Germany, e Kooperationslaboratorium Ruhrverband, Emschergenossenschaft und Lippeverband (EGLV), Essen, Germany * Correspondence: Martin D. Klassen, Institut f ¨ ur Umwelt & Energie, Technik & Analytik e.V. (IUTA), Bliersheimer Str. 58-60, 47229 Duisburg, Germany (e-mail: klassen@ iuta.de) . Ethics: not applicable (no human/animal work). The data that supports the findings of this study are available from the corresponding author, Martin D. Klassen, upon reasonable request. Supplemental digital content is available for this article. Direct URL citations appear in the printed text and are provided in the HTML and PDF versio ns of this article on the journal ’ s Web site (www.ejoncologypharmacy.com) . Copyright © 2026 the Author(s). Published by Wolters Kluwer Health, Inc. This is an open access article distributed under the terms of the Creative Commons Attribution-Non Commercial-No Derivatives License 4.0 (CCBY-NC- ND) , where it is permissible to download and share the work provided it is properly cited. The work cannot be changed in any way or used commercially without permission from the journal. The European Journal of Oncology Pharmacy (2026) 9:1(e60) Received: 15 August 2025 / Accepted: 17 December 2025 Available online 27 March 2026 http://dx.doi.org/10.1097/OP9.0000000000000060 1 mAbs rituximab, bevacizumab, and trastuzumab are classified as hazardous substances by European Chemicals Agency (ECHA) due to their potential to induce specific target-organ toxicity within the immune system after repeated exposure, [8] as well as their documented [9] or suspected reproductive toxicity. [10] Furthermore, proteinaceous surface contamination can promote microbial growth as it is a potential substrate for microorganisms to colonize the stainless steel surface which is referred to as biofouling. [11] In addition, dried mAb residues can be a potential source of particle emissions, which is undesirable in cleanroom environments. Thus, an evaluation of different cleaning strategies for mAbs can be essential to fulfill the requirements of EU GMP Annex 1 if these active ingredients are handled in the sterile manufacturing of medicinal products. The aim of this study was to investigate the cleaning efficiency of various cleaning solutions and procedures to remove mono - clonal antibodies from stainless steel surfaces. Different alkaline solutions and 70% isopropyl alcohol were evaluated as cleaning solutions. In addition, various procedures such as a single-pass wiping with one cleaning solution in comparison with different multipass procedures of subsequent wiping with an alkaline solution and isopropyl alcohol were investigated based on their cleaning efficiency. 2. Materials and Methods 2.1. Chemicals and reagents Bevacizumab (BVCZ) and trastuzumab (TTZ) were purchased as Avastin 25 mg/mL and Herceptin 150 mg (reconstituted to 20 mg/ mL) from Roche Pharma (Basel, Switzerland). Daratumumab (DRTM) was acquired as Darzalex 20 mg/mL from Janssen-Cilag (Neuss, Germany). Cetuximab (CTX) was obtained as Erbitux 5 mg/mL from Merck KGaA (Darmstadt, Germany) and omalizumab (OMLZ) as Xolair 150 mg/mL from Novartis (Basel, Switzerland). Rituximab (RTX) was purchased as Truxima 10 mg/mL from Celltrion Healthcare (Budapest, Hungary). Guanidine hydrochloride (GuHCl), ammonium bi - carbonate (NH 4 HCO 3 ), dithiothreitol (DTT), tris(hydroxy - methyl)aminomethane hydrochloride (TrisHCl), phosphate buffered saline (PBS) tablets, and formic acid (FA) were supplied by Sigma-Aldrich (St. Louis). Trypsin was acquired from Promega Corporation (Madison). Acetonitrile was purchased from Th. Geyer (Renningen, Germany). Ultrapure deionized water was produced by PURELAB Chorus 1 (Veolia Water Technologies Deutschland GmbH, Celle, Germany). 0.9% sodium chloride solution was purchased from B. Braun Melsun - gen AG (Melsungen, Germany). Sodium hydroxide was pur - chased from Fluka (Seelze, Germany) and dissolved in water as a 0.1 M NaOH solution. The sterile alkaline cleaning product perform sterile mucasol (2% Mucasol), 70% isopropyl alcohol (70% IPA) branded as perform sterile alcohol IPA and perform sterile mix dry wipes were kindly provided by Sch ¨ ulke & Mayr GmbH (Norderstedt, Germany). Kimtech Science 7557 wipes from Kimberly-Clark Inc (Neenah) were used for wipe sampling. 2.2. Cleaning solutions The sterile alkaline cleaning product perform sterile Mucasol is a concentrate which was used as 2% Mucasol solution ( v/v ) for cleaning. Perform sterile Mucasol concentrate includes tripotas - sium orthophosphate ( $ 30%) as active ingredient, in addition to less than 5% amphoteric surfactants. The pH of 2% Mucasol is approximately 12, while the pH of 0.1 M NaOH is calculated to be 13. Perform sterile Mucasol includes tripotassium orthophos - phate as an active ingredient. 70% isopropyl alcohol (70% IPA) branded as perform sterile alcohol IPA is a ready-to-use cleaning solution for cleanroom environments (sterile filtered) with 70% isopropyl alcohol as active ingredient. 2.3. Preparation of surfaces, cleaning procedures, and wipe sampling The cleaning validation studies were performed on smooth, unpolished, and deburred stainless steel substrates (type: AISI 304) with a total area of 200 cm 2 (20 3 10 cm). The steel grade used in this study (AISI 304) is comparable with the steel grade used in safety cabinets designed for the aseptic production of parenteral cytostatic preparations in accordance with DIN 12980 and DIN EN 12469. [12] First, the substrate surfaces were contaminated with defined concentrations of each biopharma - ceutical product. Two dilutions with monoclonal antibodies were prepared for this purpose. In stock solution A, the medicinal products were diluted in deionised water to antibody concentra - tions of 1.1 mg/mL, 1.1 mg/mL, 5.5 mg/mL, and 2.75 mg/mL for CTX, DRTM, RTX, and TTZ, respectively. Stock solution B was prepared with a final concentration of 16.5 mg/mL for both BVCZ and OMLZ. The test surfaces were spiked with 0.3 mL of stock solution A and 0.1 mL of stock solution B. This results in concentrations on the surface of 1,650 ng/cm 2 (CTX and DRTM), 4,125 ng/cm 2 (TTZ), and 8,250 ng/cm 2 (RTX, BVCZ, and OMLZ), respectively. The applied concentrations vary depending on the determined detection limit. The spiked substrates were left to dry at room temperature. The time between spiking and cleaning was a maximum of 1 hour. After drying, the surfaces were cleaned with different cleaning solutions and procedures according to the study design. Perform sterile mix dry wipes were used for the cleaning procedures. The wipes were weight and soaked with 300 wt% of the respective cleaning solution. The cleaning procedure was developed based on the wipe sampling method proposed by Hetzel et al. [13] In brief, the wipe was placed over the edge of the hand near the little finger. Wiping was performed systematically in overlapping lanes with constant pressure to ensure thorough surface cleaning. After completing one pass in overlapping lanes, the outer part of the cleaning cloth was folded inward, and the wiping front was cleaned in a single lane; this process is referred to as single-pass wiping and required one cleaning wipe. For multiple cleaning passes, both the direction of the lanes and the soaked cleaning cloth were changed for each pass. Multipass cleaning means the sequential application of an alkaline solution followed by an alcohol solution (two-agent), performed as either one-cycle two-agent (requires 2 wipes, one for each cleaning solution) or three-cycle two-agent (requires 6 wipes, 3 for each cleaning solution). In the three-cycle two-agent wiping procedure, the same cleaning solution was used 3 times, with the wiping direction and cleaning cloth changed after each pass. A new cleaning solution was then introduced for the next set of wiping passes. With regard to the intended application of the cleaning process on stainless steel surfaces, the study design was designed to be gentle on the material by first using alkaline solutions and then alcohol solutions. A phosphate buffered saline (PBS) buffer (10 mM phosphate, 2.7 mM potassium chloride, and 137 mM sodium chloride, pH 7.4) was produced by dissolving 5 tablets in 1 L of ultrapure deionized water. After cleaning, the wipe sampling was performed using Kimtech Science 7557 wipes and PBS buffer as sampling solution. Sampling was performed in 3 different 2 Thissen et al. European Journal of Oncology Pharmacy (2026) 9:1 directions, with each pass using a fresh wipe presoaked with 1 mL of PBS. The sampling technique described here was established by Hetzel et al. [13] and the same as the cleaning technique. All 3 sampling wipes were placed in the same sampling cup and extracted by adding 30 mL of PBS followed by manual shaking. Aliquots of the extracts were filtered through a 0.45 m m regenerated cellulose (RC) syringe filter. The substrates were cleaned after each experiment by three-cycle two-agent cleaning procedure before storage for next use. 2.4. Tryptic digest Before analysis, 80 m L of the monoclonal antibodies in the wipe sample extracts were digested using trypsin. The protocol was adapted from Reinders et al. [14] and modified to accommodate a total sample volume of 80 m L. In specific terms, this means an increase in all volumes by a factor of 1.6. The substances used and their concentrations remain unchanged. 2.5. Quantitation of the cleaning efficiency The LC-MS/MS method was adapted from Reinders et al., [14] with modifications to the calibration parameters for the analysis of wipe sample extracts. The LC-MS/MS equipment used remained the same. In brief, 60 m L of the enzymatically hydrolyzed samples were injected onto an Eclipse Plus C18 RRHD 1.8 m m, 2.1 3 50 mm column (Agilent Technologies, Waldbronn, Germany) and separated at 60 °C using a linear gradient elution from 10% B to 27.8% B over 8 minutes. Mobile phase A was water 1 0.1% formic acid, and mobile phase B was acetonitrile 1 0.1% formic acid. A specific peptide sequence was selected for each of the mAbs. The individual mAbs were quantified in positive polarity after electrospray ionization according to the following MS/MS transitions: m/z 588.3 . m/z 359.2 (BVCZ), m/z 633.8 . m/z 359.2 (CTX), m/z 532.8 . m/z 725.3 (DRTM), m/z 870.0 . m/z 359.2 (OMLZ), m/z 803.9 . m/ z 926.5 (RTX), and m/z 887.0 . m/z 359.2 (TTZ). Quantification was performed with Analyst 1.6.3 Build 5095 (AB Sciex, Darmstadt, Germany). The detailed method parameters for the LC-MS/MS analysis as well as the validation can be taken from Reinders et al. [14] The calibration curve for this study consisted of ten equidistant points with concentration ranges of 0.01 – 1 m g/mL for DRTM, 0.1 – 10 m g/mL for CTX, 0.25 – 25 m g/mL for TTZ, and 0.5 – 50 m g/ mL for BVCZ, RTX, and OMLZ. The calibration was measured daily for the individual trials and diluted in wipe sample matrix. Positive controls were included to determine the recovery for all individual mAbs, meaning that wipe samples were taken directly from spiked substrates without a cleaning step. The mean recoveries ranged between 57.2 and 101.5% with BVCZ 88.0% (95% confidence interval [95% CI]: 77.9 – 98.1%), CTX 100.7% (95% CI: 92.4 – 108.9%), DRTM 57.2% (95% CI: 17.1 – 97.2%), OMLZ 101.8% (95% CI: 96.5 – 107.2%), RTX 101.5% (95% CI: 96.8 – 106.3%), and TTZ 96.5% (95% CI: 90.6 – 102.3%). For each cleaning procedure, 4 substrates were spiked, dried, and cleaned by the individual procedure, followed by wipe sampling, preparation, enzymatic hydrolysis and LC-MS/ MS analysis. Each of the 4 samples was measured once. The cleaning efficiency was calculated based on the amount of antibody detected in the wipe samples collected after the cleaning procedures. The lowest concentration on the calibration curve was considered the technical limit of quantification (LOQ). The LOQ was 82.5 ng/cm 2 for BVCZ, OMLZ, and RTX, 41.25 ng/ cm 2 for TTZ, 16.5 ng/cm 2 for CTX, and 1.65 ng/cm 2 for DRTM. The spiking, sampling, and analysis of the stainless steel substrates were performed by a total of 3 different individuals. Owing to the amount of data (N 5 4), no statistically relevant evaluation of the operator ’ s influence can be made. Nevertheless, this approach reduced the personal influence on the analysis results. 2.6. Statistical analysis The statistical analysis was performed with OriginLab software (Origin 2021b, OriginLab, Northampton). In comparison with the total amount spiked on the stainless steel substrates, a maximum cleaning efficiency of 99% can be reached for all antibodies, except for daratumumab (99.9%), based on the technical LOQ. Thus, for better comparability in statistical analysis, the data were first scaled to the range of 0 – 100% cleaning efficiency. Note that in this study, a cleaning efficiency of 100% means that the residual amount of mAbs was below LOQ. A 2-way analysis of variance (ANOVA) with repeated measure - ments (RM) followed by a Bonferroni-corrected post hoc test was applied to the data set consisting of factor A (6 mAbs) and factor B (7 cleaning procedures) with 4 repeated measurements on each combination. For all statistical analyses, the level of statistical significance was set at a 5 0.05, resulting in 95% confidence interval (95% CI). A 1-way RM ANOVA followed by a Bonferroni-corrected post hoc test was applied to groups with significant differences according to the 2-way RM ANOVA followed by a Bonferroni-corrected post hoc test. 3. Results 3.1. Single-pass cleaning Figure 1 shows the cleaning efficiencies of different single-pass cleaning procedures for mAbs on stainless steel. In Figure 1, the alkaline cleaning solutions 2% Mucasol and 0.1 M NaOH yielded comparable mean cleaning efficiencies of 98.3% (95% CI: 96.8 – 99.9%) and 98.3% (95% CI: 96.5 – 100.1%) for all mAbs investigated, respectively. The mean Figure 1. Cleaning efficiencies of different single-pass cleaning procedures performed to remove mAbs from stainless steel. Data are presented as mean 6 95% CI with N 5 4 independent replicates per group. A cleaning efficiency of 100% corresponds to a residual amount of mAbs below LOQ. BVCZ, bevacizumab; CTX, cetuximab; DRTM, daratumumab; OMLZ, omali - zumab; RTX, rituximab; TTZ, trastuzumab. 3 Thissen et al. European Journal of Oncology Pharmacy (2026) 9:1 cleaning efficiency of 70% IPA was 57.6% (95% CI: 48.0 – 67.1%), which was significantly lower compared with the alkaline cleaning agents according to a Bonferroni-corrected post hoc test following 2-way RM ANOVA. Furthermore, significant differences in the cleaning efficiencies of various mAbs could be observed when cleaning with 70% IPA. In particular, the cleaning efficiency of 70% IPA for OMLZ is significantly lower (mean: 41.4%, 95% CI: 37.4% – 45.3%) than for all other mAbs, while DRTM (mean: 66.6%, 95% CI: 55.5 – 77.7%) results in significantly higher cleaning efficiency according to a Bonferroni-corrected post hoc test following one-way repeated measurements ANOVA performed on cleaning efficiencies of 70% IPA. 3.2. Multipass cleaning In Figure 2, the cleaning efficiencies of different multipass cleaning procedures for the removal of mAbs on stainless steel are demonstrated. Multipass cleaning refers to the sequential applica - tion of an alkaline solution (2% Mucasol or 0.1 M NaOH) followed by an alcohol solution (70% IPA), performed either once each (one-cycle) or 3 times each (three-cycle) in succession. One-cycle two-agent wiping with 2% Mucasol and 70% IPA led to a mean cleaning efficiency of 99.3% (95% CI: 98.0% – 100.5%), while one-cycle two-agent wiping with 0.1 M NaOH and 70% IPA resulted in a mean cleaning efficiency of 99.9% (95% CI: 99.8% – 100.1%). The cleaning efficiencies for the individual antibodies slightly vary for both one-cycle two-agent procedures. However, no significant differences in the removal can be observed between the mAbs. Furthermore, the comparison between both one-cycle two-agent wiping procedures revealed no statistically significant differences. Three-cycle two-agent wiping with 2% Mucasol followed by 70% IPA resulted in a mean cleaning efficiency of 99.5% (95% CI: 98.3% – 100.7%). The three-cycle two-agent wiping procedure with 0.1 M NaOH and 70% IPA yielded a mean cleaning efficiency of 100% (95% CI: 100.0 – 100.0%), which means that all mAbs were removed below LOQ (see Table S1, http://links.lww.com/ EJOP/A8 in the supporting information). However, no statistically significant difference was observed between the two three-cycle, two-agent wiping procedures. In addition, no statistically signifi - cant difference could be detected between the two-agent one-cycle and two-agent three-cycle procedures. 4. Discussion This study is, to our knowledge, the first to evaluate the cleaning efficiency of different cleaning solutions and procedures for removing various mAbs from stainless steel. While several studies focused on the cleaning or decontamination efficiency of different cleaning solutions for various cytostatic agents, [1 – 6] there is a lack of data on the decontamination of mAbs. This study successfully implemented a methodology to validate the cleaning efficiency of various cleaning solutions and procedures on mAbs deposited on stainless steel. In the first part of the study, the efficiency of single-pass procedures with either alkaline (2% Mucasol or 0.1 M NaOH) or alcohol (70% IPA) solutions was investigated. The alkaline cleaning solutions yielded significantly higher efficiencies in the removal of mAbs from stainless steel than 70% IPA, despite the different chemical composition of their active ingredients (sodium hydroxide vs. tripotassium phosphate in addition to amphoteric surfactants). According to the literature, both alkaline and alcohol cleaners lead to denaturation of proteins. However, while sodium hydroxide enhances the solubility of denatured proteins, [15] isopropyl alcohol results in fixation of proteins on stainless steel. [16] Therefore, the results found in this study are consistent with the literature. The significantly lower cleaning efficiency of 70% IPA for OMLZ might be explained by the lower isoelectric point (pI) of OMLZ (pI 7.3) [17] in comparison with other mAbs (pI 8.3 – 9.4 of BVCZ, CTX, RTX, TTZ). [18] It is assumed that due to the lower pI, OMLZ ’ s net charge is lower or approximately neutral in 70% IPA leading to a weaker solubility in the cleaning solution and stronger fixation on stainless steel, and thus to a lower cleaning efficiency. However, for DRTM, the pI value is approximately 8.4 – 9.0, [19] which is comparable with the pI of the other mAbs. Therefore, more research on this effect is necessary to fully understand the mechanisms involved in the cleaning process with 70% IPA and to explain the differences between the mAbs. In the second part of the study, the efficiency of multipass procedures was evaluated, starting with one-cycle two-agent procedures with alkaline solution (2% Mucasol or 0.1 M NaOH), followed by an alcohol solution (70% IPA). The mean cleaning efficiencies of the one-cycle two-agent wiping procedures with an alkaline followed by an alcohol solution (99.3% and 99.9%) were slightly higher than the cleaning efficiencies of the single-pass procedures with only alkaline cleaners (98.3% and 98.3%). However, these differences were not significant accord - ing to a Bonferroni-corrected post hoc test following 2-way repeated measurements ANOVA. In addition, mean cleaning efficiencies of the three-cycle two-agent wiping procedures with an alkaline followed by an alcohol solution (99.5% and 100%) are higher than the respective mean cleaning efficiencies of the one-cycle two-agent wiping procedures (99.3% and 99.9%) and single-pass procedures with alkaline cleaners (98.3% and 98.3%). However, no statistically significant differences in the mean cleaning efficiencies were detected between all procedures. Korczowska et al. [20] reported cleaning recommendations of the European Society of Oncology Pharmacy (ESOP) for pharmacies and wards handling cytotoxic drugs. Cleaning with 70% IPA on a daily basis and a two-agent cleaning with 0.05 M Figure 2. Cleaning efficiencies of different multi-pass cleaning procedures (one - cycle two-agent/three-cycle two-agent) performed to remove mAbs from stainless steel. Data are presented as mean 6 95% CI with N 5 4 independent replicates per group. A cleaning efficiency of 100% corresponds to a residual amount of mAbs below LOQ. BVCZ, bevacizumab; CTX, cetuximab; DRTM, daratumumab; OMLZ, omalizumab; RTX, rituximab; TTZ, trastuzumab. 4 Thissen et al. European Journal of Oncology Pharmacy (2026) 9:1 NaOH followed by 70% IPA once a week are recommended. The number of repetitions for the cleaning procedure was not specified. [20] Since both cytotoxic drugs and mAbs must be prepared under aseptic conditions, they are usually handled in the same work facilities. However, in accordance with the results of this study, mAbs will not be cleaned properly from stainless steel surfaces on a daily basis, when applying the recommended cleaning procedure of 70% IPA. Surface contamination with mAbs can potentially be a source of particles or be a potential substrate for microbial growth, [11] which should be considered according to the CCS outlined in EU GMP Annex 1. [7] Owing to the addition of an alkaline cleaning step, only the weekly cleaning protocol would significantly reduce the amount of mAb residues on stainless steel. Therefore, based on the results of our study, the daily protocol should additionally include an alkaline cleaning step for cleaning of mAb residues to comply with CCS. To ensure material compat - ibility with stainless steel, the alkaline solution should be applied first and then the alcohol solution as recommended by Korczowska et al. [20] for the weekly cleaning routine. In this study, mAbs were spiked on the stainless steel substrates in concentrations ranging between 1,650 and 8,250 ng/cm 2 , which is comparable with spilling a few drops of mAb solution. Korczowska et al. [20] recommended cleaning with an alkaline solution (0.05 M NaOH) and with an alcohol solution (70% IPA), after using a spill-kit to remove the solution, which is in line with the results of our study for mAb cleaning. Based on our results, the three-cycle two-agent procedure would be recommended as the mean cleaning efficiencies are highest compared with the other cleaning procedures, albeit not statistically significant. LOQ between 1.65 and 82.5 ng/cm 2 could be achieved for the different mAbs with the applied LC-MS/MS method. However, there are no risk thresholds or action limits given for occupational exposure or contamination control, e.g., in EU GMP Annex 1, [7] on both mAbs and cytostatic drugs. Therefore, no further practical implications on risk thresholds or action limits can be derived from this study. Additional research will be required to clarify this in future work. In conclusion, the cleaning efficiency of various solutions and procedures for removing dried monoclonal antibodies from stainless steel was successfully evaluated using the implemented sampling and analysis methodology. In future studies, the established methodology could be used to evaluate the cleaning efficiency of other cleaning solutions, such as various disinfec - tants or detergents, for aged residues and on additional substrates, e.g., glass or polymer materials. References 1. L ˆ e LMM, Jolivot PA, Sadou Yaye H, et al. Effectiveness of cleaning of workplace cytotoxic surface. Int Arch Occup Environ Health 2013;86(3): 333 – 341. 2. Simon N, Odou P, Decaudin B, Bonnabry P, Fleury-Souverain S. Chemical decontamination of hazardous drugs: a comparison of solution performances. Ann Work Exp Health 2020;64(2):114 – 124. 3. Simon N, Guichard N, Odou P, Decaudin B, Bonnabry P, Fleury - Souverain S. Efficiency of four solutions in removing 23 conventional antineoplastic drugs from contaminated surfaces. PLoS One 2020;15(6): e0235131. 4. Queruau Lamerie T, Nussbaumer S, D ´ ecaudin B, et al. Evaluation of decontamination efficacy of cleaning solutions on stainless steel and glass surfaces contaminated by 10 antineoplastic agents. Ann Occup Hyg 2013; 57(4):456 – 469. 5. Yuan Z, Li Q, Tang T, Zhang M, Liu Y, Liu L. Studies on the optimization of decontamination protocol for surfaces contaminated with cytotoxic drugs in PIVAS. J Oncol Pharm Pract 2023;29(7):1565 – 1573. 6. Portilha-Cunha MF, Pedro N, Arminda A, Rar L, Sam T, Santos MSF. Tackling antineoplastic drugs ’ contamination in healthcare settings: New insights on surface cleaning approaches. J Occup Environ Hyg 2025;22: 386 – 399. 7. European commission. EU Guidelines for Good Manufacturing Practice for Medicinal Products for Human and Veterinary Use. Annex 1: Manufacture of Sterile Medicinal Products; 2022. Available at: https:// health.ec.europa.eu/system/files/2022-08/20220825_gmp-an1_en_0.pdf Accessed April 22, 2025. 8. European Chemicals Agency (ECHA). Classification, Labeling and Packaging — Rituximab. Available at: https://echa.europa.eu/de/ information-on-chemicals/cl-inventory-database/-/discli/details/230463 . Accessed November 25, 2025. 9. European Chemicals Agency (ECHA). Classification, Labeling and Packaging — Trastuzumab. Available at: https://echa.europa.eu/de/ information-on-chemicals/cl-inventory-database/-/discli/notification - details/230457/903551 . Accessed November 25, 2025. 10. European Chemicals Agency (ECHA). Classification, Labeling and Packaging — Bevacizumab. Available at: https://echa.europa.eu/de/ information-on-chemicals/cl-inventory-database/-/discli/details/225843 . Accessed November 25, 2025. 11. P ´ erez H, Vargas G, Silva R. Use of nanotechnology to mitigate biofouling in stainless steel devices used in food processing, healthcare, and marine environments. Toxics 2022;10(1):35. 12. Berner International GmbH. Technical Information for CLAIRE® XL — Safety Cabinet. Available at: https://berner-safety.de/out/pictures/ ddmedia/broschure-safety-cabinet-claire-xl.pdf . Accessed December 12, 2025. 13. Hetzel T, vom Eyser C, Tuerk J, Teutenberg T, Schmidt TC. Micro-liquid chromatography mass spectrometry for the analysis of antineoplastic drugs from wipe samples. Anal Bioanal Chem. 2016;408(28):8221 – 8229. 14. Reinders LMH, Noelle D, Klassen MD, et al. Development and validation of a method for airborne monoclonal antibodies to quantify workplace exposure. J Pharm Biomed Anal 2022;221:115046. 15. Wiese H, Geißler H, Augustin W, Scholl S. Diffusive mass transfer and protein removal in the alkaline cleaning of a jellylike whey protein fouling layer. Heat Mass Transfer 2024;60(5):829 – 840. 16. Prior F, Fernie K, Renfrew A, Heneaghan G. Alcoholic fixation of blood to surgical instruments — a possible factor in the surgical transmission of CJD? J Hosp Infect 2004;58(1):78 – 80. 17. Wang Y, Zheng C, Zhuang C, et al. Characterization and pre-clinical assessment of a proposed biosimilar to its originator Omalizumab. Eur J Pharm Sci 2022;178:106292. 18. Goyon A, Excoffier M, Janin-Bussat M-C, et al. Determination of isoelectric points and relative charge variants of 23 therapeutic monoclonal antibodies. J Chromatogr B Analyt Technol Biomed Life Sci 2017;1065 – 1066:119 – 128. 19. National Library of Medicine (US). A Phase 2 Study of Daratumumab in Patients With Relapsed or Refractory Waldenstr ¨ om Macroglobulinemia. ClinicalTrials.gov. Available at: https://clinicaltrials.gov/study/ NCT03187262 . Accessed December 12, 2025. 20. Korczowska E, Crul M, Tuerk J, Meier K. Environmental contamination with cytotoxic drugs in 15 hospitals from 11 European countries — results of the MASHA project. Eur J Oncol Pharm 2020;3(2):e24. 5 Thissen et al. European Journal of Oncology Pharmacy (2026) 9:1