A non-transitory computer-readable medium storing instructions for determining an estimated fixation duration of an at least partially fixed test biological specimen, comprising: (a) obtaining test spectral data from the test biological specimen, wherein the obtained test spectral data comprises vibrational spectral data derived from at least a portion of the biological specimen, wherein the obtained test spectral data comprises an averaged vibrational spectrum derived from a plurality of normalized and corrected vibrational spectra, wherein the plurality of normalized and corrected vibrational spectra are obtained by: (i) identifying a plurality of spatial regions within the test biological specimen; (ii) acquiring a vibrational spectrum from each individual region of the plurality of identified regions; (iii) correcting the acquired vibrational spectrum from each individual region to provide a corrected vibrational spectrum for each individual region; and (iv) amplitude normalizing the corrected vibrational spectrum from each individual region to a pre-determined global maximum to provide an amplitude normalized vibrational spectrum for each region; (b) deriving fixation features from the obtained test spectral data using a trained fixation estimation engine, wherein the fixation estimation engine is trained using training spectral data sets acquired from a plurality of differentially fixed training biological specimens and wherein the training spectral data sets comprise at least class labels of known fixation durations; characterized by (c) quantitatively determining an estimated fixation duration of the at least partially fixed biological specimen based on the derived fixation features, wherein an amount of time the at least partially fixed test biological specimen has been subjected to a fixation process is predicted. (a) obtaining test spectral data from the test biological specimen, wherein the obtained test spectral data comprises vibrational spectral data derived from at least a portion of the biological specimen, wherein the obtained test spectral data comprises an averaged vibrational spectrum derived from a plurality of normalized and corrected vibrational spectra, wherein the plurality of normalized and corrected vibrational spectra are obtained by: (i) identifying a plurality of spatial regions within the test biological specimen; (ii) acquiring a vibrational spectrum from each individual region of the plurality of identified regions; (iii) correcting the acquired vibrational spectrum from each individual region to provide a corrected vibrational spectrum for each individual region; and (iv) amplitude normalizing the corrected vibrational spectrum from each individual region to a pre-determined global maximum to provide an amplitude normalized vibrational spectrum for each region; (b) deriving fixation features from the obtained test spectral data using a trained fixation estimation engine, wherein the fixation estimation engine is trained using training spectral data sets acquired from a plurality of differentially fixed training biological specimens and wherein the training spectral data sets comprise at least class labels of known fixation durations; characterized by (c) quantitatively determining an estimated fixation duration of the at least partially fixed biological specimen based on the derived fixation features, wherein an amount of time the at least partially fixed test biological specimen has been subjected to a fixation process is predicted.