Luminol and some of the footprint evidence

Part X in a series on the Knox/Sollecito case

The story surrounding the footprints in the murder of Meredith Kercher is complex, and it may take more than one post to cover it. However, the recent jury’s statement included a reference to the footprint evidence as one indicator of guilt. In this post we will summarize the analyses of both pro-prosecution and pro-defense points of view with respect to a bloody footprint found on a bathmat and the three luminol-positive footprints in the hallway.

Chemistry and forensics of luminol
Luminol was used to visualize some of the footprints. Luminol is typically used as a mixture that includes sodium carbonate and sodium perborate. Luminol reacts with the iron atom in hemoglobin, and also reacts with other substances, such as bleach and fruit juice, or with substances that contain a metal ion that can catalyze the light-producing chemistry. Fruit pulp is rich in certain peroxidase enzymes that have an iron ion that has much in common with the iron in hemoglobin. In hemoglobin and in the peroxidase family of enzymes the iron ion is bound to four nitrogen atoms of an organic molecule called protoporphyrin IX. The combination of iron and protoporphyrin IX is called heme, which is tightly bound to the protein portions of hemoglobin or catalase, respectively. A fifth nitrogen atom from a histidine residue within the protein also coordinates the iron ion (Frey and Hegedus, Enzymatic Reaction Mechanisms, pp. 203-209). Heme is called a prosthetic group, a nonprotein molecule that helps a protein to do its job.

Both hemoglobin and peroxidases are proteins, but only peroxidases are enzymes (biological catalysts). In the luminol reaction however, both hemoglobin and peroxidases are acting catalytically. A paper by Barni et al., “Forensic application of the luminol reaction as a presumptive test for latent blood detection,” Talanta 72 (2007) 896–913, is a review of luminol chemistry and forensics. Hydrogen peroxide oxidizes luminol with the production of light. The iron ion within hemoglobin or other substances is a catalyst; in other words, one molecule of hemoglobin converts many molecules of luminol and hydrogen peroxide to produce 3-aminophthalate. Thus the catalytic behavior of metal ions partially explains the sensitivity of this test. Since photons of light are emitted in this reaction, the process is classified as a chemiluminescent reaction.

This paper discusses whether one can tell whether or not the substance reacting with luminol is really blood:

“Due to the possible presence of these substances at the crime scene, the luminol test must not be considered sufficiently specific to permit an unequivocal identification of blood [15,18,51,88,89]….

Generally visual examination is used when the luminol test is employed in a forensic situation, rather than instrumental detection of the luminescence. An experienced practitioner may distinguish the true blood-catalyzed chemiluminescence from that produced by other substances by the evaluation of parameters observable to the naked eye such as emission intensity, duration and spatial distribution. However this approach may also lead to misinterpretation, due to a subjective, informal and non-quantitative evaluation, for example, because its intensity is qualitatively much weaker than that expected for blood. In other circumstances an emission of similar intensity may be thought to derive from diluted bloodstains and is accepted. Therefore, caution should be exercised when using the test.”

Thus the Luminol test is a presumptive test and should be confirmed by one that is more specific for blood. However (http://www.friendsofamanda.org/luminol.html), “The prosecution's key forensic witness told the court she can tell by looking at a luminol reaction whether it involves blood or something else, but she did not perform any scientific tests to validate this claim.”

Perugia-Shock
The prosecution attributed one hallway footprint to Raffaele Sollecito. Francesco Vinci testified that the luminol footprint in the hallway is a human footprint but one that cannot be attributed to anyone in particular (http://perugia-shock.blogspot.com/2009/09/sollecitos-expert-questions-all.html). The shape of the big toe and the print of the putative second toe were key elements to his conclusions. Frank Sfarzo wrote, “With such limited elements Vinci is only able to rule out Raffaele (or the girls) and to say that the print is compatible with Rudi. And we can only admire his honesty and consistency.”

Additional posts on the footprints by Frank Sfarzo
http://perugia-shock.blogspot.com/2009/05/footprint-is-mr-sollecitos.html
http://perugia-shock.blogspot.com/2008/06/footprints-are-amandas.html
http://perugia-shock.blogspot.com/2009/07/coroner-sarah-gino-accuses.html

Darkness Descending
Now let us turn to the analysis of the footprints that Colonel Luciano Garofano gave to the author of the book Darkness Descending, by Paul Russell and Graham Johnson (Pocket Books, 2009). Colonel Garofano is a well-known forensic scientist who recently retired from the Carabinieri. He was much friendlier to the prosecution’s case than to the defense’s case, and I do not agree with everything that he said in this book, particularly with respect to the footprint on the blue bathmat discussed below.

“The other problem I have is the way the Luminol was applied. The size of the blobs shows that it was not carefully vaporized but squirted. That creates two problems. It dilutes the sample and it dilates the print. We have a print attributed to Sollecito, which matches his foot in the size of the big toe, the width of the metatarsus and the width of the heel, but does not present the characteristic details each of our feet present. The print can be said to be compatible, but not 100 per cent.”

“Now let’s have a look at the prints attributed to Amanda Knox. There’s one in her bedroom facing the exit to the room, and there are two right feet in the corridor walking in the direction of the victim’s room. The same goes here. The method of application of the Luminol is insufficiently subtle to positively identify a foot, but the result can be said to be generally compatible with Amanda Knox’s.

“But I didn’t see who else they compared the prints with. Just Rudy, Amanda, and Raffaele? So we only have a choice between those? We don’t have the footprint of other women or men, as a comparison? Pity.”

Colonel Garofano believed that it is likely that Raffale’s foot made the print on the blue bathmat, but he did not sound certain. However, he did not discuss the fatal criticism that the prosecution’s expert witness made a serious error in the measurement of this print, discussed below. Colonel Garofano did not address the fact that Rudy’s big toe looks nothing like Raffaele’s, and the bathmat print. Nor did he seriously entertain the possibility that the prints are not even blood. He also did not explain why the print attributed to Ms. Knox points toward Meredith’s room and why there is not a full set of prints. These are serious omissions in that it is difficult to picture how the prints could have been made during the commission of the murder.

Science Spheres
Mark Waterbury discussed some of the problems in the footprint evidence in the hallway and elsewhere (http://www.sciencespheres.com/2009/10/methods-of-polizia-pseudoscientificaa.html):
“Luminol glowing footprints were found in a hallway, and some may have been Amanda's, it is hard to know for sure because they were only compared with her feet, and found to be ‘compatible.’ Again, no controls. Meredith, Laura, Filomena, none of the other resident's feet were compared to these footprints. The footprints were tested for blood, and it came out negative. No blood. So, why are they important? Amanda lived there, after all.”

“Amanda's DNA was said to be found in one of these footprints. Did they also test a meter away from the footprints, to see if her DNA was all over the apartment where she lived? No. That would have been another control experiment. Was the DNA actually associated with the footprint, or did it just happen to be there, because the resident's DNA was all over their apartment, as people's DNA usually is? We will never know. They skipped the control experiments, and presented results without any reference.”

With respect to Meredith’s DNA and the luminol-enhanced footprints, I would argue that not finding her DNA is evidence against the proposition that the substance is her blood. However, finding her DNA would be not conclusive evidence that the substance is her blood. That is because there could be non-blood derived biological material from Meredith mixed into the footprint. In other words finding her DNA would have been consistent with blood being responsible for the luminol reaction but would not demand that conclusion.

Perugia Murder File
A powerpoint, Dear-Mr-Marriott-I-Shrunk-the-Black-Kid.pps, authored by Kermit (http://www.perugiamurderfile.org/viewtopic.php?f=5&t=36) attempted to rebut arguments presented at the Friends of Amanda website. The first of these is that the prosecution’s expert witness Mr. Rinaldi, made a serious error in his size measurements. Kermit’s presentation also addresses two of the problems in ascribing the bloody footprint to Mr. Sollecito, the narrowness of the big toe (http://www.friendsofamanda.org/footprint.html) and the appearance of a mark near the big toe. Mr. Guede’s big toe is shorter than his second toe, but Mr. Sollecito’s second toe does not even show up in his footprint. Mr. Guede’s big toe is narrower than Mr. Sollecito’s. According to Kermit, the reason that the bloody imprint of the big toe seems narrower than Mr. Sollecito’s is that the raised tufts of the blue bathmat have picked up more blood than the unraised portions. Likewise, the mark Sollecito’s expert witness Francesco Vinci would ascribe to a second toe (http://perugia-shock.blogspot.com/2009/09/sollecitos-expert-questions-all.html), Kermit claims is Mr. Sollecito’s big toe. Kermit said elsewhere that his reason for writing his powerpoint presentations were to refute the lone wolf theory of the crime, because the prints were of varying sizes.

I do not find Kermit’s arguments about the raised tufts to be convincing. There is a raised tuft of lighter color in between the big toe and the second mark, possibly from Guede’s second toe. If Kermit were correct, that portion of the bathmat should be as dark as the rest. The validity of Kermit’s measurements has also been questioned by two commenters at a forum at James Randi Educational Foundation (http://forums.randi.org/showthread.php?t=161229&page=126).

Defense-friendly blogs
Charlie Wilkes of the website Friends of Amanda presented the argument that the bathmat footprint measurements made by the prosecution’s witness, Lorenzo Rinaldi, contained a critical error. (http://www.friendsofamanda.org/footprint_measurements.html). Mr. Wilkes also discussed the footprints elsewhere (http://knoxarchive.wordpress.com/2009/11/27/manuela-comodi-asks-for-common-sense/). The site Injustice in Perugia gives a detailed account with images of the footprints (http://www.injusticeinperugia.org/footprints.html). These two sites make several telling points, especially with respect to the luminol-positive footprints in the hallway. The footprint L9 (in the knoxarchive numbering scheme) is attributed to Amanda by Rinaldi. Yet is alone, and it points toward, not away from Meredith’s room. L6 and L7 are two right feet, one of which is attributed to Raffaele. The footprint in the hallway attributed to Raffaele look quite indistinct to me. The other one is attributed to no one at all, and one wonders to whom it belongs. None of the footprints tested positive for blood and none were positive for Meredith’s DNA.

None of the three footprints were part of a trail. It is difficult to see how Amanda Knox and Raffaele Sollecito could have removed other footprints (as part of an alleged cleanup) without also cleaning up Rudy Guede’s nearby bloody shoeprints.

Prosecutorial tunnel vision
The failure to obtain reference footprints is disturbingly reminiscent of the lack of reference DNA from Laura or Filomena, the other two flatmates. It suggests that once the investigators locked onto Amanda and Raffaele, they did not reopen their field of vision. Crime journalist Mario Spezi said about the word “compatible,” in Douglas Preston’s book, The Monster of Florence, which covers a serial murderer in that city.

“Compatible, not compatible, and incompatible are the baroque inventions of Italian experts who don't want to take responsibility. Using 'compatible' is a way to avoid admitting they haven't understood anything. Was the bullet in Pacciani's garden inserted into the monster's pistol? 'It is compatible.' Was that laryngeal break inflicted by someone who intended to kill? 'It is compatible.' Was that painting done by a monstrous psychopath? 'It is compatible.'”

“Perhaps yes, perhaps no--in short, we don't know! If the experts are chosen by the investigators, they say their results are 'compatible' with the theories of the prosecution; if they are chosen by the defendants they say that their results are 'compatible' with the theories of the defense. That adjective should be outlawed!”

Conclusions
Luminol was overapplied to the footprints in the hallway, and a full set of reference footprints was not taken. These two points alone make the prosecution’s attribution of two the footprints to Ms. Knox and Mr. Sollecito, respectively, impossible to accept as solid conclusions. The unattributed footprint forces one to ask who else was in the hallway and when. Moreover, the luminol-visualized footprints did not contain Meredith’s DNA and cannot be said to be blood. The failure to obtain a positive test for blood is possibly the most serious problem for the prosecution’s case. Although I do not claim to be an expert in the analysis of footprints, the bathmat footprint looks like a better match to Rudy Guede than to Raffaele Sollecito. The footprints in the hallway may date from a time prior to the murder, or on the following morning. The lack of reference footprints is one more indication that the forensics in this case was not pursued in an entirely objective manner.