## 4 Discussion
### Limitations  
The present study design has several confounders and limitations. First of all, the study considers the standard 5-ALA dose of 20 mg/kg. Although toxicological safety has been demonstrated for higher doses of 5-ALA [24], it is unknown whether higher doses of 5-ALA would provide more fluorescence in meningioma. The present study will not allow the generation of data to this end, which might be the subject of follow-up trials.  
The study relies on study surgeons actively interrogating areas in which they would consider additional information from fluorescence to be helpful. This study does not force surgeons to perform biopsies, except for biopsies from bulk tumor, because mandating biopsies from any intracranial tissues would be unethical. Theoretically, a surgeon could perform the entire procedure without interrogating tissue once, except for the bulk tumor. This inherent uncertainty regarding the number of biopsies and, respectively, the productive participation of surgeons can only be resolved by the careful education of surgeons about study procedures and study aims.  
The study is designed as an all-comer study, irrespective of meningioma grade, anatomical location, size, or urgency of surgery. Since the obligatory assessment of each tissue interrogation/biopsy is reviewed by external panelists and their assessments are based on videos, video quality must be high. For deep-seated tumors, such as petroclival meningiomas, which are operated under high magnifications, optimal conditions for obtaining evaluable videos, specifically under blue light, are difficult to achieve. Retraction for the sake of better video imaging would not be considered acceptable for obvious reasons. It is likely that if patients with such deep-seated tumors are recruited, the number of biopsies qualifying for evaluation will be low even if there is useful fluorescence.  
Furthermore, inclusion in this study is based on pre-operative MR imaging of a meningioma. With minor exceptions, the pre-operative assumption will be correct. However, in rare cases, other pathologies might mimic meningioma and result in erroneous recruitment [25; 26], such as carcinoma metastasis, dura based malignant gliomas, lymphomas, plasmocytomas, undifferentiated sarcomas, chondrosarcoma, or schwannomas. Such patients would be included in the safety population but would obviously not qualify for the prespecified endpoints. Regarding the different grades of meningioma, it is difficult to pre-operatively reliably predict histological differentiation. Past experience has suggested that fluorescence accumulates independently of meningioma grade [9]. Due to the relative paucity of high-grade meningiomas compared to grade I meningiomas, the number of malignant subtypes in this study will be small.  
In some centers, meningiomas are embolized preoperatively in order to reduce blood loss and to reduce the complexity of surgery [27]. We hypothesize that due to reduced blood flow to tumors or necrosis of tumor tissue, drug delivery to the tumor would be impaired. Thus, such patients are excluded from this study, and it will be unknown whether any benefit from FGR extends to embolized tumors.  
Finally, this study relies on perfectly functioning video documentation. We have encountered operations in which the microscope videos did not work sufficiently. Under these circumstances, panel review cannot occur. Data from such patients will be included in the safety population and in separate exploratory evaluations of data based on the surgeons' impressions only but will be excluded from analyses based on the Biopsy Efficacy Analysis Population. To compensate for these patients, the number of patients enrolled in the study has been slightly expanded.