## 4 Molecular Analyses
### Protein Analysis and Applications  
The study of proteins, or proteomics, is a powerful tool for the study of a parchment material, as it can provide not only a sensitive approach to characterize the protein composition of a given sample, but also to identify the species of the proteins themselves and thus, the protein-based material that was used to manufacture a specific object [38]. This molecular approach is gradually becoming a recognized method in archaeological research, as one of the major advantages of this technology is that it requires a small sample and it is less laborious and costly than DNA analysis is. This approach includes basic protein analyses or more in-depth proteomic analyses to identify the additional proteins on the surface of the parchment material [6]. Specifically, in the study of ancient parchment, a basic protein analysis has been successfully applied to determine the species of animal that was used in the parchment's manufacture. This method is called peptide mass fingerprinting (PMF) and it uses enzymatic digestion to cleave the proteins at specific amino acid sites, thereby forming a peptide mixture. After the collection has been performed, each sample is chemically treated to allow for the extraction of collagen from the skin. Although all of animal skin contains collagen, the mixture of peptides in the collagen of each species is unique. Using mass spectrometry techniques, the molecular weights of the individual peptides in each sample can be determined, thereby allowing a peptide mass fingerprint (PMF) to be generated for each of them. The fingerprint of one particular protein that has been obtained using mass spectrometry can identify the species of the animal that it derived from [38]. Until recently, the protein analysis of parchment required its destructive sampling, which consequently severely limited the number of analyses that could be performed on a given document or on several documents in an archive. As an example, in the study by [PERSON] et al. [39] on the 13 th century \"Marco Polo Bible\", the authors needed 5 mg of single-leaf parchment to identify that it was made of calfskin by the analysis of the peptide sequences. Thus, the same authors pointed out that the study was biased because more sheets could not be analyzed due to the destructive technique that was used and that it could have been made from different types of parchment, depending on what material was available at the time of its production. A further study by [PERSON] et al. [38] also showed the use of destructive sampling techniques to identify a Qur'an folio that is dated to the 9 th c. as being made of sheep skin. Authors have also analyzed and compared some other objects that are made of animal skins, and they have stated that in all of the objects that were investigated, the ages and sample conditions had no significant effect on the resulting protein spectra and their subsequent identification. In order to avoid destructive techniques being used in the study of ancient manuscripts, [PERSON] et al. [7] went a step forward,  
Figure 4: Sampling of an ancient parchment manuscript using PVC erasers by gently rubbing the frame of the manuscript. using the non-invasive sampling method that is based on PVC polymer rubbers, which has the additional advantage of preserving the proteins in the PVC polymer residues at room temperature and without the need for the special storage of them. These authors performed a comparative analysis of the same sample using conventional MS techniques that require destructive sampling to occur, as well as the non-destructive electrostatic MS using the PVC eraser sampling method. They obtained the same, if not better, results with the PVC sampling than they did when they were using a real fragment of parchment.