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Analysis of Normal Blood & Menstrual Blood found on crime scene

Menstrual blood contains a variety of substances, such as:1. Blood: Menstrual blood is a mixture of uterine lining blood and blood from tiny blood vessels that burst during menstruation. 2. Endometrial Tissue: The endometrium, or uterine lining, sheds during menstruation and contributes significantly to menstrual blood. 3. Cervical Mucus: By assisting in the removal of menstrual waste from the body, cervical mucus also aids in menstrual discharge. 4. Vaginal Secretions: Vaginal secretions can mingle with menstrual blood, changing the consistency and makeup of the latter. 5. Cells: Epithelial cells, red and white blood cells are present in the menstrual blood. 6. Water: The majority of menstrual blood is water, which helps keep it liquid.

Analysis of menstrual blood and normal blood found in crime scene
The Normal blood contains red blood cells (RBCs) which carries haemoglobin, that transports oxygen from the lungs to the body. The white blood cells (WBCs) are necessary to the immune system and aid in the fight against illnesses and infections. Platelets are in charge of making the blood clot. When you are hurt, they assist in stopping the bleeding. Plasma is the liquid part of the blood, which is around 55% of the total blood. It consists nutrients including water, glucose, electrolytes, proteins (like albumin & globulins), hormones and waste products. Some additional products which includes waste materials (urea, creatinine, etc.) and electrolytes (sodium, potassium, calcium, etc.). Several tests can be performed to examine normal blood discovered at a crime scene. These examinations aid in identifying, describing, and maybe connecting the blood to specific people. Among the important tests are:
1. Presumptive Blood Test: Based on the presence of haemoglobin, a preliminary test is done to determine whether a substance is blood, such as the Kastle-Meyer test.
2. Blood Typing: Blood samples are examined to determine the Rh factor (positive or negative) and the ABO blood group (A, B, AB, or O).
3. DNA Analysis: A blood sample can be used to extract DNA, which can then be studied using procedures like PCR and STR analysis. This makes individual profiling possible.
4. Serology: This refers to examinations for the presence of serum and plasma proteins, as well as other blood-related components, which can reveal further details about the blood sample.
5. Hematological analysis: Blood samples may be studied under a microscope to determine a variety of features, including the quantity of red, white, and platelet cells in the blood.
6. Blood Enzyme Testing: Blood enzyme tests can be used to identify or characterize the blood by recognizing specific enzymes that are present in the blood.
Tests to analyse and Identify blood found in the crime scene
Presumptive tests: It is analytical chemical based method based on peroxidase enzyme activity on the heme portion of the hematin present in blood stains.
1. Benzidine Test (Alder and Alder 1904):
Because haemoglobin in our blood has peroxidase activity, it can catalyse the breakdown of a peroxide, such as hydrogen peroxide, and liberate an oxygen molecule. This oxygen molecule then combines with the test reagent, such as benzidine to produce a noticeable colour shift.
Materials required are Benzidine solution (1.5gms), Glacial acetic acid (13ml), re-distilled water (57ml) forming hydrogen peroxide and the sample (suspected blood stain). It involves the reaction of benzidine with haemoglobin, resulting in a characteristic blue-green colour change.
2. Kastle Mayer Test: The Kastle-Meyer test is a cost-effective and swift method employed in crime scene investigations to detect the existence of blood. When phenolphthalein interacts with hydrogen peroxide in the presence of haemoglobin, it undergoes a colour change from clear to pink.
Materials required include the Kastle Mayer reagent (2gms), Stock solution of Phenolpthalein (20gms), Potassium Hydroxide (100ml), Distilled water. A swab of the sample is taken which is placed into a test tube which contains approximately 1cm of distilled of H2O, swirl to release the contents. Several drops of Kastle/Meyer reagent, followed by several drops of H2O2. The positive outcome will show a bright pink colouration.
3. Luminol Test: The luminol test is a forensic technique used to detect the bloodstains which are invisible to the naked eye. Luminol reacts with the iron in haemoglobin, producing a blue glow in low-light conditions. This test is highly sensitive and can identify blood even after attempts to clean or conceal it. However, it does not specify whether the blood is human or animal. The results are temporary, so photographs or documentation are crucial for later analysis.
This method includes the reagent preparation with the help sodium carbonate, sodium percarbonate, minopthal hydrazide and distilled water.
This test is mainly used for the detection of Menstrual blood: -
1. The SERATEC PMB Test, a device specifically geared towards detecting D-dimer, a marker for menstrual blood, was developed on the basis of the SERATEC HemDirect test, intended to detect human haemoglobin and hence the presence of blood. This ground-breaking duplex assay separates samples containing menstrual fluid among a variety of sample types, including dried, liquid menstrual & peripheral blood along with samples from diverse ages and genders. It also accurately identifies human haemoglobin. When compared to other menstrual blood detection techniques such microscopic examination, Lugol’s Iodine Staining Method, and Dane’s Staining Method, the SERATEC PMB Test performed better in terms of sensitivity and accuracy. In particular, it provided benefits in terms of speed, repeatability, and simplicity of usage. The membrane of the reacting SERATEC PMB Test card and the sample in the buffer were found to be valuable sources of DNA for extraction and typing, enhancing its utility for forensic purposes.
Confirmatory tests for blood analysis:
1. Teichmann test: The materials used for blood group detection include chemical reagents like teichmann reagent (consisting of KCl 0.1%, KI 0.1%, and Glacial acetic acid of 10ml), Hayem’s fluid, Turk’s solution, distilled water, Sahli’s Haemoglobinometer, Hemacytometer grid, HCl and antiserum ABH.
Sample collection involved selecting 20 normal female subjects aged 18-25 years and collecting both the liquid and stained menstrual blood samples in containers and in airtight or zip-locked plastic envelopes respectively. Additionally, approximately 2ml of normal blood was also collected from the same individuals.
The procedure includes distinguishing between menstrual & normal blood samples of aforementioned individuals. Haemoglobin levels were determined using Sahli’s haemoglobinometer, where anticoagulated blood was mixed with 0.1 N HCL and left for 5 to 7 minutes to form haematin acid. The colour of acid haematin was compared with a calibration tube solution, and distilled water was added to the acid haematin until the colours matched & readings were noted. Furthermore, RBC and WBC cell counts were obtained using Neubauer’s Hemacytometer slide for both samples from the same individual. Finally, an attempt was made to determine the blood group of the menstrual blood samples with the help of antisera ABH.
2. Crystal Test: -
Formation of crystals by haemoglobin derivatives like haematin, haemin etc. with different salts in acidic and alkaline aqueous medium.
It has been demonstrated that separating menstrual fluid from peripheral blood using these techniques is quite difficult, and applying more sophisticated molecular approaches calls for a significant level of expertise. Furthermore, a thorough forensic validation is required because results could be impacted by intra- and inter-individual variances across donors.

Himika Ghosh

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