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Haemotaphonomy. The "strange" world of bloodstains' cells.
Haemotaphonomy
The "strange"
world of
bloodstains' cells
by Policarp Hortolà
Haemotaphonomy or hemotaphonomy
refers
to "the
study of bloodstains, and especially of the changes in appearance and size of the
cellular components, as well as the characteristics of their cell position and
appearance in function of the superficial topography and composition of the
substrate".
This term was proposed by the author
in a paper published in 1992 in
Forensic Science
International.
The presence of all kind of
residues on implements agrees with the criminalistic well-known Locard’s
Principle of Exchange ('every contact leaves traces'). On the other hand, actuopalaeontology and experimental archaeology are both based upon the Lyell’s
Principle of Actualism ('the present is the key to the past'). A short-time
preservation of specimens is a sine qua non precondition to do feasible a
(palaeobiological, bioarchaeological, forensic) longer one.
Vertebrate blood (i.e., blood sensu
stricto) is a cell suspension into a fluid medium
(the plasma). There are three types
of blood cells: erythrocytes
(red blood cells, RBCs), leukocytes (white blood cells, WBCs)
and thrombocytes (platelets, in mammals). Unlike the other vertebrates
(cyclostomes, fishes, amphibians, reptiles, and birds), mammals have
anucleate RBCs (akaryocytes or erythroplastids). As an
exception to mammals, the salamanders belonging to the family Plethodontidae
have some proportion of erythroplastids, with the California slender salamander
Batrachoseps attenuatus possessing nearly 95%
anucleate erythrocytes. Also, the pearlside teleostean fish Maurolicus
muelleri has akaryocytes. Due to the lack of
nucleus, the typical mammalian erythrocytes are shaped as
biconcave discs (discocytes). This does not apply to the family Camelidae, where
RBCs are oval (ovalocytes). Other physiological shapes, which are
minor or pathologic, are: echinocytes (burr or berry cells), dacryocytes (tear
drop cells), schizocytes (helmet cells), keratocytes (horn cells), drepanocytes
(sickle cells), and many others.
The occurrence of (at least
morphological) preservation
of anucleate,
mammalian RBCs
in
bloodstains has been
reported
even in Oldowan tools from Sterkfontein Cave (South Africa), assigned to be ca.
2 Myr old. These corpuscles have also been identified in prehistoric immovable items, such as
an early Holocene building at Çayönü Tepesi (Turkey), containing anucleate RBCs, human
immunoglobulin G (IgG) and both human
and non-human haemoglobin (Hb) on a stone slab.
The transfer activities for non-human prehistoric bloodstains on stone tools are
centred in game hunting and butchery activities. The potential for human blood
smears encompasses a wider range of items. These would include stone tool
manufacturing, assaults, surgery, rituals (burial dismemberment, sacrifice,
mutilation, scarification), and –whether
ritual or not– cannibalism. From
the practical point of view, a supplementary interest of bloostains is that they can be dated electron spin
resonance, a non-destructive technique that requires less than 0.25 g of sample
and its dating range could cover all of the Quaternary times.
The study of the different RBC and
plasma-matrix morphologies exhibited in bloodstains represents a field with implications
in
forensics,
prehistoric archaeology,
palaeoanthropology, etc.
In forensic analysis, the presence of RBCs in a smear is considered a blood
confirmation. Despite this fact and the ancient RBC evidence, interest in bloodstain analysis has been
largely focused on the molecular level, and the knowledge of the morphological characteristics of bloodstain-origin
RBC had not
been taken into account.
Using scanning electron microscopy (SEM), the author has carried out a new
approach to the study of bloodstains: the haemotaphonomy of
mammalian RBCs. Materials encompass
human and non-human blood, lithic and non-lithic, hard and soft substrate, and
from 1 month to more than 10 years stains' ageing.
The largest part of the found smear-origin RBC
shapes shared morphology with those described in haematology. But two
time-independent RBC shapes were interpreted as due specifically to blood drying
phenomena. Thus, they may be considered as genuine RBC morphologies
characteristic of mammalian bloodstains, obviously not found under physiological
conditions:
Moon-like shapes, which would be due to
erythrocyte-plasma interaction when drying, and
Negative replicas, that would be related to
imprinting by dried plasma matrix.
This long-term research led to the author to
propose both a terminology and
a systematics for the smear-origin erythrocytes, which would be
valid for, at least, mammalian
blood.
Taphoerythrocytes.
Three-month-old
smear on stone. Author's blood.
Hecatocytes.
Six-month-old smear
on stone. Blood from collared peccary
(Tayassu tajacu).
Janocytes.
About-eight-month-old smear
on urban asphalt. Human-suspected blood.
Physiocytes.
More-than-ten-year-old smear
on stone. Human blood.
TAPHOERYTHROCYTES
(from the Greek taphos, burial)
Smear-origin erythrocytes
PHYSIOCYTES
(from the Greek physis, nature)
Physiological shapes
DISCOCYTES, ECHINOCYTES,
SPHEROCYTES,
etc.
Biconcave cells, burr cells,
spherical cells, etc.
KELIDOCYTES
(from the Greek kelida, smear)
Smear-only shapes
HECATOCYTES
(from the Greek Hecate, the Greek supreme goddess identified in
heaven with Selene, moon the deity)
Moon-like shapes
JANOCYTES
(from the Latin Ianus, Janus the Roman double-faced god)
Negative replicas
Systematics for the smear-origin
erythrocytes, wich is valid for, at least, mammalian
blood.
Author's papers on
haemotaphonomy
(full-text pdf available upon request)
Keywords:
Scanning electron microscopy, blood smears, erythrocytes, organic residues,
prehistoric archaeology, actualistic palaeoanthropology, forensic biology.
SEM analysis of red blood cells
in aged human bloodstains. Forensic Science International
vol. 55, pp. 139–159,
1992.
Mammal red blood cells (RBC) in bloodstains have been
previously detected by light microscopy on stone tools from as early as
100,000±25,000 year ago. In order to evaluate the degree of morphological
preservation of erythrocytes in bloodstains, an accidental human blood smear
on white chert and several experimental bloodstains on hard substrate (the
same stone
-
white chert; another type of stone
-
graywacke; a non-stone support
-
stainless steel) where stored in a room, in non-sterile and fluctuating
conditions, for lenghths of time ranging from 8 to 18 months. Afterwards,
the specimens were coated with gold and examined by a Cambridge Stereoscan
120 scanning electron microscope. Results revealed a high preservation of
RBC integrity, with the maintenance of several discocytary shapes, a low
tendency to echinocytosys and a frequent appearance of a moon-like
erythrocytary shape in the thinner areas of the bloodstains.
SEM characterization of blood stains on
stone tools. The Microscope vol. 40 (2), pp. 111–113
[Editor's ‘Errata’ in vol. 40 (3), p. vi],
1992.
Mammalian red blood cells (RBC) in bloodstains have been
previously detected by light microscopy on stone tools circa 100,000
years old. To observe and characterize the bloodstain-original RBC, a modern
replica of ancient blood residues was examined by scanning electron microscopy
(SEM). To simulate a Stone Age process a collared peccary (Tayassu tajacu)
cadaver was skinned using a paleolithic-like white chert knife, which was then
smeared with blood plus serous liquid. After drying in the open air for one
week, the tool was stored at unsterile and fluctuant room conditions. After six
months, a detached fragment of the bloodstain on a SEM stub was coated
with gold and examined at an accelerating voltage of 15 kV using a Cambridge
STEREOSCAN 120 scanning electron microscope. Results reveal protuberant
moon-like shapes which are interpreted to be characteristic of RBC.
Application of SEM to the study of red blood
cells in forensic bloodstains. Microscopy and
Analysis vol. 40,
pp. 19–21 (UK)
& vol. 28, pp.
21–23
(EU), 1994.
Although in forensic analysis the presence of red blood cells in a smear is
considered to be a confirmation of blood, to date morphological researches using
electron microscopy dealing with superficial preservation or imprints
characterisation of erythrocytes in bloodstains have not been carried out.
Short-time preservation of specimens is a "sine qua non" precondition to
bioarchaeological or palaeobiological preservation.
So a part of author’s researches on ancient
blood residues has been focused on developing a methodology to study forensic
suspected bloodstains – considering as 'forensic' all modern blood smears of
unknown origin– covering substrates from hard to (absorbent) soft ones.
In this article dealing with
'urban' forensic hemotaphonomy, field and scanning electron microscopy
procedures, as well as showing examples of achieved results on either soft
(paper handkerchief) and hard (urban asphalt on a crossing-walk) smear
substrates are reported.
Experimental SEM
determination of game mammalian bloodstains on stone tools.
Environmental Archaeology vol. 6, pp. 99–104,
2001.
The presence of
morphologically complete mammalian red blood cells (RBC) from bloodstains
has been previously evidenced in prehistoric implements. While the presence
of ancient non-human blood on a prehistoric tool is an evidence of the real
use of this on an animal resource, the presence of RBC in a smear is an
evidence as being blood. In a simulation of a prehistoric predation human
operative chain, mammalian bloodstains on palaeolithic-like chert implements
were obtained from two specimens belonging to order Artiodactyla: collared
peccary (Tayassu tajacu, family Tayassuidae) and Dorcas gazelle (Gazella
dorcas, family Bovidae). After one year, the unburied peccary blood
smear and the buried gazelle one were coated with gold and then examined by
a scanning electron microscope. Results revealed the presence of preserved
RBC with several shapes as those found in haematological studies, as well as
moon-like shapes and negative replicas, two
bloodstain-characteristic morphologies which are interpreted as due,
respectively, to erythrocyte-plasma interaction when drying and to
imprinting by dried plasma matrix.
Morphological characterisation
of red blood cells in human bloodstains on stone: a systematical SEM study.
Anthropologie vol. 39, pp. 235–240,
2001.
Mammalian erythrocytes or red blood cells (RBC) have been
previously reported as forming part of residues on prehistoric implements.
On the basis of the Principle of Actualism, several thick smears of human
blood were obtained on chert.
After increasing lengths of storage time span (1‑36 months), the bloodstains
were micromorphologically studied via scanning electron microscopy. Results
revealed, in all the smears, the presence of an erythrocyte acme‑zone with
packed RBC, as well as negative replicas and moon‑like shapes. Morphologies
were found to be time‑independent, and furthermore those erythrocyte
acme‑zones with packed RBC to be thick‑bloodstain characteristic.
Red blood cell haemotaphonomy of
experimental human bloodstains on techno-prehistoric lithic raw materials.
Journal of Archaeological Science vol. 29, pp. 733–739,
2002.
Mammalian
erythrocytes or red blood cells (RBC) -whose presence in a
smear is a blood evidence- have been previously reported as forming part of
residues on prehistoric implements assigned to be as far as around two
million years old. On the basis of the Principle of Actualism, bloodstains
from human individuals were obtained on obsidian, limestone and chert, and
then stored in a unsterile room under microclimatically unmanipulated
fluctuating conditions, for lengths of time ranging from 7 years, 6 months
to 10 years, 2 months. Afterwards, the bloodstains were doubly coated with
carbon and gold and then examined by a JEOL JSM-6400 scanning electron
microscope (SEM). Results revealed a high preservation of erythrocyte
integrity, with several shapes as those found under physiological conditions
and a significant presence of moon-like shapes plus a minor one of negative
replicas, two morphologies that are interpreted as specifically related to
blood drying phenomena. These results agree with several previously reported
SEM analyses of younger mammalian bloodstains on chert and materials other
than obsidian and limestone, and lead to consider the moon-like shapes
(hecatocytes, a term ex novo) and negative replicas (janocytes, another term
ex novo) as the genuine RBC morphologies characteristic of (at least
mammalian) bloodstains.
The “strange” world of bloodstain
cells. A brief overview of haemotaphonomy.
Problems of Forensic Sciences
vol. 57, pp. 16–23, 2004.
Mammals are
the only vertebrates that have anucleate red blood cells (RBC's). In this
zoological class, RBC's typically have the shape of biconcave discs. The
cytomorphological study of RBCs in bloodstains is an issue with implications
in fields such as forensic biology or prehistoric archaeology. Using scanning
electron microscopy, the author has pioneered a new approach to the study of
bloodstains, which has led moreover to a general terminology and systematics
for smear-origin mammalian RBC's. This paper summarises the results of more
than 10 years of research in this field, by presenting the main morphological
features of mammalian erythrocytes, when in smears.
SEM examination of human erythrocytes in
uncoated bloodstains on stone: use of conventional as environmental-like SEM
in a soft biological tissue (and hard inorganic material).
Journal of Microscopy
vol. 218, pp. 94–103, 2005.
Although nowadays the so-called
environmental scanning electron microscopes (ESEMs) allow the observation of
the samples without metal or carbon coating, many conventional scanning
electron microscopes (SEMs) are still in use. On the other hand, the
presence of erythrocytes (red blood cells, RBCs) in a smear is considered a
blood confirmation. Such a presence has been previously reported even in
Lower Stone Age implements. In previous works, I have reported several
studies dealing with cytomorphology of RBCs in bloodstains using scanning
electron microscopy with standard specimen preparation procedures, i.e. via
coating the samples before SEM analysis. In order to explore the potential
of conventional SEM as environmental-like SEM in haemotaphonomical studies,
two alkaline (limestone) and two acid (flint) rock fragments were smeared
with human blood from a male and a female. The bloodstains obtained in this
way were then air dried indoors and stored into a non-hermetic plastic box.
Afterwards, the smears and their rock substrates were examined directly
without coating, via secondary electrons, using a JEOL JSM-6400 scanning
electron microscope. Satisfactory results reveal the capability of a
conventional SEM to work in secondary-electron mode as an environmental-like
SEM on these kinds of biological and inorganic materials, and probably in
many other biological and non-biological samples.
Using an SEM as an ESEM to study minute
human bloodstains on stainless steel.
Microscopy and
Analysis vol. 20(6) [new series],
pp. 15–17
(UK), pp. 5–7
(EU), pp. 23–25
(US) & pp. 11–13
(AP), 2006.
Because many conventional,
high-vacuum scanning electron microscopes (SEMs) are still in use, their
full potential should always be explored. With this aim in mind, two
uncoated stainless-steel blood lancets, used for finger puncturing in a
study carried out 20 months earlier, were examined for possible blood smears
in an SEM using secondary electron imaging at an accelerating voltage of 0.5
kV. Minute bloodstains and some of their erythrocytes were found. As was
previously revealed in uncoated bloodstains on stone, this study clearly
demonstrated that a conventional SEM can be used in secondaryelectron mode
just like an environmental SEM to examine these biological materials on a
stainless-steel substrate.
Secondary-electron SEM bioimaging of
human erythrocytes in bloodstains on high-carbon steel substrate without
specimen preparation.
Micron vol. 39, pp. 53–55, 2008.
The imaging of most biological
samples via conventional scanning electron microscope (SEM) in
secondary-electron mode involves routinely some kind of specimen
preparation. Conventional SEMs are still used when a low-vacuum or
variable-pressure SEM (usually known as ‘environmental’, or ESEM) is not
available. But that preparatory approach may be undesirable in certain
cases, for instance in museum specimens, forensic evidences or clinical
samples. This report details a simple, low-cost, and sample-saving
bioimaging protocol without specimen preparation, by using removable plastic
conducting carbon cement, and then working under ex-profeso SEM conditions,
i.e., by using an SEM in secondary-electron mode just like an ESEM. The
successful use to image up to high magnifications human erythrocytes in
bloodstains on an extensively bloodsmeared, high-carbon steel surgical blade
is reported as an example of the potential of this procedure.
Author's details
Policarp Hortolà
graduated in
biological sciences and then did a doctoral programme
on the sedimentary record
and palaeoenvironmental evolution at the
University of Barcelona. Later on, he
was awarded with a pre-doctorate fellowship
in order to research within the
Atapuerca Research Team.
He received his PhD degree at the
Rovira i Virgili University.
His doctoral thesis on the morphology of mammalian erythrocytes
in bloodstains, with a prehistorical bias, was awarded with the
'Doctorate Special Prize'
of this university.
He is currently working at
the Rovira i Virgili University as
an Investigador
Ordinari (Senior Researcher & Lecturer). He
teaches both human palaeocology and
molecular archaeology in the
European Master's Degree in Quaternary Archaeology and Human Evolution.
He also
taught epistemology during two academic years.
His biographical record is listed in Marquis
Who’s Who in the World
and Who's Who in Science and Engineering.
Author's address
Dr. Policarp Hortolà
Area of Prehistory
(CSIC
Associate Research Unit)
Rovira i Virgili University
Plaça de la Imperial Tàrraco 1
E-43005 Tarragona
Catalonia - Spain (E.U.)
E-mail:
policarp.hortola@urv.cat
policarp@prehistoria.urv.cat
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