Using bright field microscopy, the cotton fibres of 27 black garments could be classified into 7 groups. To investigate the discriminating power of isotope ratio mass spectrometry (IRMS), the largest of the 7 groups consisting of 11 garment samples was analysed using IRMS. The discriminatory power of the IRMS technique using a combination of carbon, hydrogen and oxygen isotope ratios was greater than 96%, only two pairs of samples out of the possible 55 pairing combinations could not be discriminated. However, these two pairs of samples which could not be distinguished by IRMS could be readily distinguished using fluorescence microscopy. Thus the combination of optical microscopy and isotope ratio analysis provided a powerful method for discrimination of black cotton fibres.