Every year, 1.7 million woman are diagnosed with breast cancer worldwide. Breast Conserving Surgery (BCS) – where not the whole breast but only the tumour is dissected – is the most common and preferable treatment for these woman. However, 20 to 55% of the patients’ needs a second surgery because of positive margins. A dissected tumour has a positive margin if it lacks a border of 2mm healthy tissue at any point. Currently, no instruments are available to provide intraoperative distinction between adipose and muscle tissue with real-time feedback. This thesis evaluates a ‘smart’ electrosurgical knife, which uses Diffuse Reflectance Spectroscopy (DRS) to discriminate between tumour and adipose tissue to assess margins intraoperative with real-time feedback. Previous research has shown that DRS can discriminate between tumour and adipose tissue. In addition, electrosurgical cutting in adipose tissue does not affect the measured DRS spectra. This thesis focuses on detecting muscle tissue under a layer of adipose tissue with DRS while cutting the adipose tissue using an electrosurgical knife. An answer to this research question is provided based on four experiments. All experiments are performed on pork muscle- and adipose tissue. DR spectra are measured with multiple probes and a ‘smart’ electrosurgical knife. PNSas software is used to acquire the spectra and fit the data. First, the influence of electrosurgical cutting and coagulation is described. This is accomplished by examining the shape and size of the cut and measuring DR spectra. Second, the transition from adipose to muscle tissue was identified in the resulting DR spectra. In addition, chromophore concentrations are retrieved from the fit data. The second experiment was repeated on cut muscle tissue. Moreover, the DR spectra obtained while cutting with an electrosurgical knife are presented.