Hematology Finals PDF

HEMATOLOGY MLS 413 | LECTURE | PRELIMS INTRODUCTION TO CLINICAL HEMATOLOGY HISTORY OF CLINICAL HEMATOLOGY Bizzozero challenged contemporary concepts Athanasius Kircher (1657) involving leukocyte...

HEMATOLOGY MLS 413 | LECTURE | PRELIMS INTRODUCTION TO CLINICAL HEMATOLOGY HISTORY OF CLINICAL HEMATOLOGY Bizzozero challenged contemporary concepts Athanasius Kircher (1657) involving leukocytes in blood coagulation but Antonie van Leeuwenhoek (1674) concluded that participation of blood platelets and Giulio Bizzozero (late 1800s) white cells in fibrin formation was conceivable. James Homer Wright (1902) Bizzozero investigated the formation of blood cells ATHANASIUS KIRCHER and described and named platelets, the small 1646 using a microscope: plague victims particles important in clotting. He is often recognized o 1646 used microscope to study the blood of as having rediscovered Helicobacter pylori, a plague victims bacteria that causes chronic gastritis disease. 1658 Scrutinium Pestis: "little worms" or JAMES HOMER WRIGHT “animalcules" in the blood Pathology Laboratory at the Massachusetts General o 1658 Scrutinium Pestis noted the presence of Hospital “little worms” or “animalcules” in the blood and 1902 modification of Romanowsky stain concluded that the disease was caused by Megakaryocyte origin of platelets (Boylston Medical microorganisms. Prize in 1908) o The conclusion was correct although it is likely He is remembered eponymously by the blood cell that what he saw were in fact red or white blood stain that bears his name. Wright’s stain facilitates cells and not the causative agent. differentiation of blood cell types. He was the proposed hygienic measures: isolation, quarantine, recipient of the Gross prize in 1905 for his burning clothes worn by the infected and wearing publication on actinomycosis and the Boylston facemasks Medical Prize in 1908 for his discovery of the origin o He proposed hygienic measures to prevent the of platelets spread of disease such as isolation, quarantine, Romanowsky stains are neutral stains composed of burning clothes worn by the infected and a mixture of oxidized methylene blue (azure) dyes wearing facemasks to prevent the inhalation of and Eosin Y. The azures are basic dyes that bind germs acid nuclei and result in a blue to purple color. The ANTONIE VAN LEEUWENHOEK acid dye, eosin, is attracted to the alkaline Leeuwenhoek made microscopes consisting of a cytoplasm, producing red coloration. single high-quality lens of very short focal length; at Wright's stain is a hematologic stain that facilitates the time, such simple microscopes were preferable the differentiation of blood cell types. It is classically to the compound microscope. a mixture of eosin (red) and methylene blue dyes. It In 1674 he likely observed protozoa for the first time is used primarily to stain peripheral blood smears, and several years later bacteria. urine samples, and bone marrow aspirates, which o Those “very little animalcules” he was able to are examined under a light microscope. In isolate from different sources, such as rainwater, cytogenetics, it is used to stain chromosomes to pond and well water, and the human mouth and facilitate diagnosis of syndromes and diseases. intestine. He also calculated their sizes. It is named for James Homer Wright, who devised GIULIO BIZZOZERO the stain, a modification of the Romanowsky stain, in Discovered: 1902. Because it distinguishes easily between blood o H. pylori cells, it became widely used for performing o Function of platelets differential white blood cell counts, which are Platelets were discovered by the Italian pathologist routinely ordered when conditions such as infection Giulio Bizzozero in 1882. He observed them or leukemia are suspected. microscopically in the circulating blood of living HEMATOLOGY animals and in the blood removed from the blood haima + logos vessels. BLOOD He discovered and carefully described blood platelet nutritive fluid function in flowing conditions and the relationship It is a connective tissue. Plasma is its ground between platelet adhesion to an artificial surface, substance. aggregation and subsequent fibrin formation and participates in the physiologic and pathologic deposition on activated platelet membrane. activities of the body RMT - 2024 1 HEMATOLOGY MLS 413 | LECTURE | PRELIMS INTRODUCTION TO CLINICAL HEMATOLOGY o Arterial blood [right]– brick red (due to increase the platelets group together to create a clot. The oxygen concentration) clot forms a scab, which stops the bleeding and o Venous blood [left]– dark red (due to decrease helps protect the wound from infection. oxygen concentration) CHARACTERISTICS OF BLOOD Fluid in vivo – inside the body, it is fluid. Red in color – due to the presence of hemoglobin (its pigment is caused by the iron component). Thick and viscous – blood is a solution. There are solutes and solvents [water]. Due to the increased number of solutes [cells, proteins, ions, or lipids], it When we get arterial blood, we usually collect in the causes the blood to appear viscous. radial artery or in the brachial artery. The angle of o ANALOGY: Three sachets of coffee in a cup insertion in arterial puncture is higher compared to cause high viscosity. venous which is lower. In venous puncture, we Slightly alkaline [pH 7.35 – 7.45] usually collect in the antecubital area from either of S.G. 1.045 – 1.065 – in further strengthening, it has the three veins (basilic, median-cubital, or cephalic). lots of solutes FUNCTIONS OF BLOOD 7 – 8% of the total body weight Respiration – delivers oxygen to organs that need Total volume: [M = 5-6 L; F = 4-5 L] oxygen, removing the carbon dioxide to the lungs. COMPOSITION OF BLOOD Nutrition – blood is a mixture that consists of solute LIQUID (PLASMA/SERUM) and solvent. In your solvent, there are a lot of Plasma is devoid with any dissolved components. There are a lot of proteins, cellular component. If you minerals, and vitamins needed for the body’s move the blood cells in one development and growth. area, it will become the Excretion – there are lot of toxins and waste buffy coat and factory cells. products produced in cells. The blood facilitates the The remaining supernatants removal of those toxins. are called plasma or serum. Homeostasis – to maintain the optimal condition of The difference of your the body plasma and serum is that the latter has no fibrinogen Body protection – due to the presence of the WBCs, because it is converted into fibrin clots. It is already it kills pathogens or microorganisms that harm the clotted. The blood in the serum is clotted then we body centrifuged it [supernatant serum]. The former is Transport – transport of nutrients and hormones mixed with an anticoagulant. In the case of o Transporting oxygen and nutrients to the lungs hematology, it is either EDTA (Ethylenediamine and tissues. tetraacetic acid) or citrate. o Forming blood clots to prevent excess blood Water [91.5%] – solvent loss. Chemicals o Carrying cells and antibodies that fight infection. o Proteins [7%] o Bringing waste products to the kidneys and liver, o Others [1.5%] which filter and clean the blood. ▪ Electrolytes o Regulating body temperature. ▪ NPN o Supplying oxygen to cells and tissues ▪ Hormones & enzymes o Providing essential nutrients to cells, such as ▪ Food materials amino acids, fatty acids, and glucose Plasma, also called blood plasma, the liquid portion o Removing waste materials, such as carbon of blood. dioxide, urea, and lactic acid Plasma serves as a transport medium for delivering o Protecting the body from diseases, infections, nutrients to the cells of the various organs of the and foreign bodies through the action of white body and for transporting waste products derived blood cells from cellular metabolism to the kidneys, liver, and o Regulating body temperature lungs for excretion. o The platelets in blood enable the clotting, or coagulation, of blood. When bleeding occurs, RMT - 2024 2 HEMATOLOGY MLS 413 | LECTURE | PRELIMS INTRODUCTION TO CLINICAL HEMATOLOGY It is also a transport system for blood cells, and it Basophil – nucleus is rarely seen because of large plays a critical role in maintaining normal blood granules and intense stain pressure. Monocyte – large indented monocytic nucleus. It is Plasma helps to distribute heat throughout the body the immature form of your neutrophil and to maintain homeostasis, or biological stability, 10 SERVICES OFFERED BY HEMA AND including acid-base balance in the blood and body. HEMOSTASIS LAB SOLID/CELLULAR COMPONENTS Specimen collection & prep’n for exam RBC Quantitative manual & instrumental measurements o 45% of blood volume of cells ▪ If centrifuged, the percent of your RBCs is Measurements of cell volumes called hematocrit after centrifugation Evaluation of cellular contents & components process. That is an indicative of total red Cellular identification blood cells population. Identification of reactive or neoplastic alterations of 12 o 4.5 – 6.5 x 10 /L cell populations WBC – can be seen in the buffy coat along with your Evaluation of leukocytes, erythrocytes and platelets. WBC can granular or agranular. platelet function 9 o 4 – 11 x 10 /L Evaluation of cellular development and formation o Granular & Agranular (BM) Platelets – are not necessarily cells. They are just Evaluation of hemostatic function merely cytoplasmic fragments of your COMMON PREFIXES USED IN THE VOCABULARY megakaryocyte that aid or facilitate during clotting. OF HEMATOLOGY o 150 – 350 x 109/L a-/an- GRANULOCYTES & AGRANULOCYTES o Lack, without Granulocytes ▪ E.g., aplasia [a – absence; plasia – o Basophils [0.5 – 1%] → Mast cells synthesis, meaning there is no production] ▪ In charge for allergies and acute immune aplastic responses o Absent, decreased o Eosinophils [2 – 4%] – in charge for sense of aniso- parasites and regulating allergic reactions. o Unequal o Neutrophils [60 – 70%] – first line of defense of ▪ E.g., anisocytosis – aniso [unequal], cyto bacteria and other foreign bodies. [cell], unequal size of cell Agranulocytes o Dissimilar o Lymphocytes [20 – 25%] – T and B cells for ante- humoral immunity o before ▪ T cells ▪ E.g., antecedent, antepartum ▪ B cells Brady- o Monocytes [3 – 8%] → Macrophages o Slow ▪ To increase the amount of phagocytosis ▪ E.g., bradycardia – slow heartrate Cyto- o Cell ▪ E.g., Cytoplasm, cytokinesis Dia- o Through ▪ E.g., diapedesis dys- o abnormal o difficult, bad ▪ E.g., dysmenorrhea [menorrhea – menstruation] – abnormal menstruation Eosinophil – large eosinophilic granules Neutrophil – appears to four nuclei erythro- RMT - 2024 3 HEMATOLOGY MLS 413 | LECTURE | PRELIMS INTRODUCTION TO CLINICAL HEMATOLOGY o red ▪ E.g., poikilocytosis – increase red blood ▪ E.g., erythrocyte cells of any shapes ferr- schis- o iron o split ▪ E.g., ferrous, ferric ▪ E.g., schistocyte – split of cells hemo- scler- o pertaining to blood o hard ▪ E.g., hemolysis, hemoconcentration ▪ E.g., sclerosis, sclerocholangitis hyper- spleen- o above, beyond o spleen o extreme thromb(o) hypo- o clot, thrombus o beneath, under ▪ E.g., thrombosis – too much clots o deficient, decreased xanth- iso- o yellow o equal, alike, same ▪ E.g., xanthelasma, xanthochromic leuko COMMON SUFFIXES USED IN THE VOCABULARY o white OF HEMATOLOGY macro- -blast o large o Primitive precursor o long -cyte mal- o Cell o bad, abnormal -ectomy removal ▪ E.g., malabsorption o Excision mega- o cut out o large, giant ▪ E.g., vasectomy meta- -emia o after, next o Blood ▪ E.g., metamorphosis -itis o change o Inflammation Mono- -lysis o one o destruction, dissolving Morph- -(o)logy o Shape o study of ▪ E.g., morphology -oma myel(o) o swelling o from the BM o tumor o spinal cord ▪ E.g., leiomyoma [leio – smooth] – smooth pan- muscle tumor o all, overall -opathy ▪ E.g., pandemic o Disease o all-inclusive ▪ E.g., cardiomyopathy [cardio – heart; myo – phleb- muscle] – there is disease in the heart o vein muscle ▪ E.g., phlebotomy -osis phago- o state, condition o eat o increase o ingest ▪ E.g., panmyelosis ▪ E.g., phagocytosis -penia poikilo- o decrease o varied, irregular o lack of RMT - 2024 4 HEMATOLOGY MLS 413 | LECTURE | PRELIMS INTRODUCTION TO CLINICAL HEMATOLOGY ▪ E.g., neutropenia, leukopenia -phil(ic) o attracted to o affinity for -plasia o cell production or repair -poiesis o cell production o formation, development -poietin o stimulates production -stasis o same o standing still -trophy o nourishment EXAMPLES OF HEMATOLOGIC TERMS Anisocytosis An + iso + cyt + osis Aplasia A + plasia no synthesis Anemia An + emia Dysmyelopoiesis Dys + myelo + poiesis dys – wrong; myelo – bone marrow; poiesis - formation Panmyelosis Pan + myel(o) + osis pan – all; myelo – bone marrow; osis – increase dysmyelopoiesis – uncountable SUMMARY History Services offered by Hematology Laboratory Terms used in the vocabulary of Hematology RMT - 2024 5 HEMATOLOGY BSMLS 3F MLS 413 | LECTURE | PRELIMS 2022 - 2023 BLOOD COLLECTION OBJECTIVES The correct order of draw in the case: Light blue top, Standard precautions on the collection of blood SST, Green top, and Lavender top. specimens. SAFETY IN THE HEMATOLOGY LABORATORY List collection equipment STANDARD PRECAUTIONS: Blood Collection Correlate tube stopper color with additive o Following the standard precautions is important. Selection of certain veins for venipuncture o All blood samples are potentially infectious with Steps recommended by CLSI: Venipuncture, order of blood-borne pathogens draw “POTENTIALLY INFECTIOUS”: Bloodborne Complications encountered in blood collection and pathogens. These standard precautions can be the proper response applied through: Good specimen quality o Handwashing Specimen rejection o PPE List reasons for inability to obtain a blood specimen o Proper disposal of sharps and infectious wastes CASE STUDY RESPONSIBILITIES OF A PHLEBOTOMIST IN CASE 1 INFECTION CONTROL A phlebotomist asks an outpatient, “Are you Susan As MedTechs, we may potentially infect others Jones?” After the patient answers yes, the because of the nature of our profession. phlebotomist proceeds by labeling the tubes and Potentially infected people: due to constant drawing the blood. What is wrong with this scenario? interaction with patients and staff. Patient identification would be more accurate if the o Observance of infection control and isolation information came from the patient themselves. This policies can be done by directly asking the patient: What is o Violations of policies be reported your name? Then the patient would answer with their A must-to-do: full name. o Good Personal health and hygiene Labeling the tubes prior collection should be avoided o Follow SP (standard precautions) at all times to prevent errors and mislabeling. Preferably, tubes PHYSIOLOGIC FACTORS AFFECTING TEST should be labeled after blood collection. RESULTS CASE 2 A patient must have blood drawn for a complete blood count (CBC), potassium level, prothrombin time (PT), and type and screen. The phlebotomist draws blood into the following tubes in this order: 1. Serum separation tube 2. Light blue (PT) 3. Lavender top (CBC) 4. Green top (potassium) Blood tests may be affected by different factors. Therefore, determination of these factors is important Question: Which of the results will be affected by the in the pre-analytical phase. incorrect order of draw? Explain. For the patient, we must first identify their Order of draw must be followed by all phlebotomists. demographic data and other physiologic factors like… This is done to avoid additive carryover which can PRE-ANALYTICAL (PEDDSS) produce erroneous results. 1. Posture (supine or erect) BCNHES 2. Exercise o Blood Culture – Yellow top 3. Diurnal rhythms o Coagulation Tube – Light blue top 4. Diet (fasting/none) o Non-additive or Serum Separation Tube – Red or 5. Stress Gold Top 6. Smoking o Heparinize Tube – Green Top As for the blood sample, we must collect the o EDTA – Lavender Top appropriate specimen in the appropriate tube. o Sodium Fluoride – Gray Top RMT - 2024 1 HEMATOLOGY BSMLS 3F MLS 413 | LECTURE | PRELIMS 2022 - 2023 BLOOD COLLECTION Examine the blood specimen whether it has been hemolyzed or contaminated. While the sample is being brought to the laboratory for analysis, observe proper storage and transportation requirements. Should adhere to the specific schedule for timed specimen collections [those tests that require fasting (FBS) or multiple collection (OGTT)] and accurately record the time of collection VENIPUNCTURE REVIEW VENIpuncture = VEIN In blood collection, there must be proper patient Venipuncture – to puncture a vein identification Three important steps prior to Adequate blood volume collection performing venipuncture: Observe the proper timing of blood collection 1. Prepare the materials Observe the proper order of draw 2. Review the min. acceptable USE OF WRONG COLLECTION TUBE volume of blood for an > 9 types of blood collection tubes individual assay/s. Each lab test requires a specific tube for blood ▪ Determine the blood volume that is required collection according to the number of tests. Each tube have different additives that is used for a 3. Determine the min. acceptable volume of blood specific laboratory test. for each type of collection tube. REMEMBER Each tube should be collected in a specific “Order of Draw” A properly collected blood samples is essential to o To avoid test errors due to additive carryover. quality laboratory outcome which may produce a domino effect towards the post-analytical phase. Strict adherence to the rules of specimen collection for the accuracy of these test results. Pre-analytical errors are major potential sources of errors Anticoagulated blood: Most common used in ORDER OF DRAW hematology (usually contained in a lavender top) Blood culture Preanalytical error (62%) Coagulation o Error in test order N (Gold) o Patient preparation Heparin fault EDTA o Patient Sodium F. misidentification EQUIPMENTS FOR VENIPUNCTURE o Specimen collection error o Tube-filling error Tourniquet o Hemolysis Gloves o Inappropriate storage Vacutainers o Transportation error Syringes Analytical (15%) Alcohol swabs o Unrecognized analytical inaccuracy Plaster o Instrument error Sharps bin Postanalytical (22%) GLOVES o Inappropriate result interpretation Prior to blood collection, we must consider whether our patient is allergic to latex or rubber. Latex RMT - 2024 2 HEMATOLOGY BSMLS 3F MLS 413 | LECTURE | PRELIMS 2022 - 2023 BLOOD COLLECTION Vinyl Ex: Glass or silica particles which activate factor 12 Nitrile [red box] in the coagulation cascade. It may also TOURNIQUET contain thrombin or activated factor 2 [orange box] Disposable elastic strap, a which converts fibrinogen to fibrin. heavier Velco strap, or a blood GEL SEPARATOR pressure cuff, latex-free Contained in a yellow top tube Applied 3 to 4 inches above Inert material that undergoes a the venipuncture site temporary change in viscosity Left on no longer than 1 during centrifugation process minute before the Enable it to serve as a separation venipuncture barrier between serum or plasma o Hemoconcentration – if left more than 1 minute and cells COLLECTION TUBES ANTICOAGULANTS Evacuated Tube System 1. EDTA (Ethylenediamine tetraacetic acid) - (ETS) Contained in a tube with a lavender top o Evacuated tube 2. CITRATE (light blue top) (plastic or glass) 3. OXALATE (gray top) o Needle The first 3 anticoagulants mentioned (EDTA, citrate, o Adapter oxalate), all chelate or bind to Calcium to prevent OSHA Recommendation: Use of plastic tubes coagulation: needed for clotting by forming insoluble whenever possible to prevent breakages calcium salts Glass tubes: (+) silicone = decrease hemolysis, prevent blood from adhering to the sides of the tube. The picture on the right shows the ETS According to the type of tube used for blood collection, a different form of the liquid component of blood may be obtained. Plasma – with fibrinogen Serum – no fibrinogen Fibrinogen is used up during blood clotting or coagulation. ADDITIVES IN COLLECTION TUBES CLOT ACTIVATORS Serum testing: takes 30-60 mins. to clot o Clotting is sped As you can see from the diagram above, calcium salts up with the use of contribute to the coagulation cascade by forming a clot activator. insoluble calcium salts and by chelating calcium, no Usually contained in clotting occurs tubes 4. HEPARIN (green top) Purposes Binds to antithrombin in the plasma 1. Accelerate o by binding to antithrombin, heparin clotting process accelerates the inhibition of thrombin and 2. Decrease activated factor ten (factor X) specimen preparation time RMT - 2024 3 HEMATOLOGY BSMLS 3F MLS 413 | LECTURE | PRELIMS 2022 - 2023 BLOOD COLLECTION Not for HEMA: interfere with WRIGHT stain = blue b/g o Heparin gives a blue background. Therefore, the identification of cells would be inaccurate or more difficult ANTIGLYCOLYTIC AGENTS 1. SODIUM FLUORIDE Inhibits glucose metabolism by blood cells, (glucose level), delayed This additive is usually used when glucose determination is needed (image above) in this diagram, sodium fluoride binds to enolase to inhibit glycolysis and red blood cells. Therefore, the glucose level in the blood is preserved RMT - 2024 4 HEMATOLOGY BSMLS 3F MLS 413 | LECTURE | PRELIMS 2022 - 2023 BLOOD COLLECTION BLOOD TUBE ORDER OF DRAW REFERENCE GUIDE COLOR TESTS SPECIAL NOTES INVERT TUBE GENTLY THIS # OF TIMES VARIES - Follow facility’s protocol for 4 drawing/labeling Blood Cultures - Draw aerobic then anaerobic - Usually, 2 separate sets from 2 O different sites R BLUE - Discard this tube N/A - Purpose of waste tube is to fill Waste Tube (if cultures are tubing with blood if cultures not drawn) weren’t drawn to allow second blue tube to fill completely BLUE - Always use a waste tube first if 4 PT, PTT, INR, Fibrinogen, D- cultures weren’t drawn Dimer, Coagulation Studies -Tube must be full RED Drug monitoring, 5 Biochemistry, Immunology GOLD 5 BMP, CMP, Hepatitis tests GREEN BMP, CMP, Troponin, 8 Lipids, Liver Panel, Ammonia (on ice) LAVENDER 8 CBC, Sedimentation Rate, Hemoglobin A1C PINK Type & Screen, Type & - Follow facility’s protocol for 8 Crossmatch, RH, Antibody drawing/labeling Screen - Send to blood bank GRAY Lactate (Lactic Acid) 8 (on ice) RMT - 2024 5 HEMATOLOGY BSMLS 3F MLS 413 | LECTURE | PRELIMS 2022 - 2023 BLOOD COLLECTION TWO-WAY NEEDLE Another method of venipuncture aside from using the syringe is by using the two-way needle in ETS Usually used in ETS with adapter Sterile, variety of lengths, and gauges (bore or opening size) Needle gauge number is inversely related to bore size: (inversely proportional) o Ex. A 23G needle has a smaller bore size than a 19G needle (Image above) Different blood collection methods Drawing blood: 19 to 23G Needle and syringe system MC needle size (adult): 21G (1 in long) Vacuum extraction system or the ETS BUTTERFLY NEEDLE Winged butterfly system (vacuum extraction) Short needle with plastic wings which may be attached to an adapter as in ETS or connected to thin tubing attached to a syringe Gauge of butterfly needles are Winged butterfly system (syringe) usually smaller CLINICAL LABORATORY STANDARDS INSITUTE Collecting blood samples: (CLSI) o Children Formerly called National Commission of Clinical o Difficult to draw blood Laboratory Standards o Geriatrics Determines, approves, and publishes the “order of o Tiny, fragile, rolling veins draw” SYRINGE Same “order of draw” is used for: Syringe method for o Glass and plastic venous blood collection tubes venipuncture is the o Evacuated system and syringe most commonly used method COLLECTION SYSTEM There are two major collection systems: N & S (needle and syringe) o Open system o Manual manipulation of plunger o Manual transfer of specimen Vacutainer System o Closed system o Quick & easy fill, no manipulation needed o Auto & direct transfer of specimen o Generally, it is safer to have or to use a vacutainer system RMT - 2024 6 HEMATOLOGY BSMLS 3F MLS 413 | LECTURE | PRELIMS 2022 - 2023 BLOOD COLLECTION TABLE 2.3 RECOMMENDED ORDER OF DRAW FOR PLASTIC VACUUM TUBES Order of use Type of tube/ usual color Additive Mode of Action Uses 1 Blood culture bottle (yellow- Broth mixture Preserves viability of Microbiology – aerobes, black striped tubes) microorganisms anaerobes, fungi 2 Non-additive tube 3 Coagulation tube (light blue Sodium citrate Forms calcium salts to Coagulation tests top) remove calcium (protime and prothrombin time), requires full draw 4 Clot activator (red top) Clot activator Blood clots, and the serum Chemistries, is separated by immunology and centrifugation serology, blood bank (cross-match) 5 Serum separator tube (red- None Contains a gel at the bottom Chemistries, fray tiger top or gold) to separate blood from immunology and serum on centrifugation serology 6 Sodium heparin (dark green Sodium heparin or Inactivates thrombin and For lithium level, use top) lithium heparin thromboplastin sodium heparin, for ammonia level use either 7 PST (light green top) Lithium heparin Anticoagulants with lithium, Chemistries anticoagulant and a gel separates plasma with PST separator get at bottom of tube 8 EDTA (purple top) EDTA Forms calcium salts to Hematology, blood bank remove calcium (cross-match) requires full draw 9 Blood tube (Pale yellow top) Acid-citrate-dextrose Complement inactivation HLA tissue typing, (ACD, ACDA, or ACDB) paternity testing, DNA studies 10 Oxalate/fluoride (light grey Sodium fluoride and Antiglycolytic agent Glucoses, requires full top) potassium oxalate preserves glucose up to five draw (may cause days haemolysis if short draw) NOTE! Since The winged butterfly system contains air in its tubing, a non-additive or discard tube should be used first to flush out the air along with some amount of blood The non-additive tube has a red top Then after using a non-additive or discard tube, the next tube used would only draw blood without any air (Image above) Table of all the tubes containing different additive and the required number of inversions 3 POSSIBLE SOURCES 1. Venous blood o Obtained through venipuncture o Syringe method o Vacuum method (ETS) RMT - 2024 7 HEMATOLOGY BSMLS 3F MLS 413 | LECTURE | PRELIMS 2022 - 2023 BLOOD COLLECTION o Winged or butterfly method a narrower diameter than the arm vein. So, there is a 2. Capillary blood potential risk of hand injury. Therefore, a smaller 3. Arterial blood gauge needle like the one in the winged butterfly SELECTION OF VEIN system and small volume tubes should be used To select a vein for venipuncture, Hand or wrist veins: we usually use the median cubital o May be used when vein. It is on the opposite side of the Median Cubital or elbow Cephalic veins are And the area (red box) is called the unsuitable or antecubital fossa unavailable o The clinician must use ORDER OF VEINS extra care to anchor 1. Median cubital vein them 2. Cephalic vein o Have a narrow diameter therefore it may be 3. Basilic vein necessary to use a small gauge needle and small So why do we usually volume evacuation tubes use the median cubital o Note: when drawing tubes from the hand, the use vein? of a butterfly apparatus can be more effective and The Median Cubital less painful. o Large o Well-anchored o Least painful o Least likely to bruise The Cephalic vein is the 2nd choice because: o It is not as well anchored o It is more painful when punctured than the Median Cubital vein Vein to Avoid – Basilic vein o Third and last choice in the antecubital fossa Other options include the jugular, scalp, and femoral o It is near the brachial artery veins. However, extra caution must be practiced o Possible artery puncture because these vessels are near important arteries o Nerves can be damaged SAFE PUNCTURE o It is located near to a vein. Therefore, there may be potential nerve damage We must practice safe puncture by avoiding: o Hitting a nerve o Puncturing arteries o Excessive or blind probing/shooting with a needle If the arm veins are quite difficult to locate or palpate, Be aware of site selection when collecting blood we can then select veins from the hand: samples However, please take note that the bones of the hand are nearer to the skin surface and the hand vein have RMT - 2024 8 HEMATOLOGY BSMLS 3F MLS 413 | LECTURE | PRELIMS 2022 - 2023 BLOOD COLLECTION VENIPUNCTURE PROCEDURE REVIEW! Select the site for venipuncture, we usually go for the Prior to blood collection, the patient’s identity should easiest vein to locate. So, preferably the median be confirmed properly by asking for their pertinent cubital vein. If the veins are not easily visible or are information easily palpated, we can apply warm compress to These information must come from the patients make veins more visible themselves Apply a tourniquet about 3-4 inches above the Sometimes a patient may bring with them a laboratory venipuncture site or 4 to 5 finger widths above the site request along with all the test required by their physician When checking for the test request, we must also assess whether the patient is ready. Ex. There is a request for FBS, we must ask the patient whether they have fasted for the appropriate amount of time 6. Ask the patient to form a fist so that the veins are more prominent. 7. Put on well-fitting, non-sterile gloves. 8. Disinfect the site using 70% isopropyl alcohol for 30 seconds and allow to dry completely (30 seconds) 9. Anchor the vein by holding the patient’s arm and placing thumb BELOW the venipuncture site. 10. Enter the vein swiftly at a 30-degree angle. 11. Once sufficient blood has been collected, release the torniquet BEFORE withdrawing the needle. To release pressure on the vein and to avoid mass or First, assemble the equipment. And then practice blood spillage. hand hygiene by washing or using alcohol Do proper patient identification and preparation. We should establish good rapport with the patient like greeting them properly and informing them of the procedure and why it is needed. This is a visual representation of venipuncture do the needle is inserted through the skin and it approaches the lumen of the vein. RMT - 2024 9 HEMATOLOGY BSMLS 3F MLS 413 | LECTURE | PRELIMS 2022 - 2023 BLOOD COLLECTION E. Needle too near vein valve – reposition or redirect the needle F. Collapsed vein – seen in dehydrated patients; prior to blood collection, we must choose good or patent veins. VENIPUNCTURE IN SPECIAL SITUATIONS AVOID SITES WITH: Prepare dry cotton and withdraw the needle gently. Once the needle is removed, apply gentle pressure over the side with cotton and then discard the used [From Left to Right] Burns, Scars, or On skins with needle and syringe in the sharp’s container. Label the Overlying Ink like Tattoos specimen after blood collection. In labelling, we put the patient’s full name, ID number, date and time of collection, and the phlebotomist’s initials. [From Left to Right] Visible Skin Lesions like If there are cases of breakages, we must discard Abrasions, Edematous or Swollen Areas or Limbs, them appropriately. Place contaminated material into Sites with Open Wounds, and Sites with Bruises and the infectious waste’s bin. Hematoma 12. Withdraw the needle gently and then give the patient Edema is an abnormal accumulation of fluid in the a clean gauze or dry cotton-wool ball to apply to the site intercellular space of the body with gentle pressure Do not perform a blood draw above an IV site 13. Discard the used needle and syringe or blood- o Specimen will become dilute with IV fluid sampling device into a puncture-resistant container. 14. Check the label and forms for accuracy. Use another arm or another site 15. Discard sharps and broken glass into the sharp’s container. Place items that can drip blood or body fluids into the infectious waste. 16. Remove gloves and place them in the general waste. Perform hand hygiene. If using soap and water, dry hands with single-use towels. Instead of drawing blood from a limb with an attached IV line, we can obtain a sample from the contralateral or opposite arm DIALYSIS PATIENTS Potential problems when it comes to the following: o Blood collection o Frequency of blood testing o Limited vein access Different Scenarios during Venipuncture NEVER be drawn from attached: A. Correct needle position – bevel up and inserted into o Cannula – is a temporary access to the patient’s the vein lumen blood in the form of a needle B. Needle inserted through vein – this is called through o Fistula – is a permanent surgical fusion of an and through and can cause a hematoma. artery and a vein C. Partial needle insertion – to remedy this, we should Preferred venipuncture site: advance the needle slightly once backflow is seen. o Hand vein D. Bevel resting on vein wall – no backflow of blood. o Vein away from the fistula and underside of the This happens when the blood might stop flowing into the arm syringe. Therefore, it is important to check the position of NOTE: ensure that dialysis pxs DO NOT BLEED (heparin the arm or hand. therapy to prevent thrombosis) - A special precaution RMT - 2024 10 HEMATOLOGY BSMLS 3F MLS 413 | LECTURE | PRELIMS 2022 - 2023 BLOOD COLLECTION should be taken to ensure that these patients do not TOURNIQUET REMOVAL bleed. Always remove the tourniquet prior to the removal POST-MASTECTOMY of the needle Patients who had a previous surgical procedure like NOTE: Failure to remove the tourniquet before mastectomy or breast removal. During a mastectomy, withdrawing the needle maintains pressure on the lymph nodes at the same side of the breast tissue are vein and causes blood to flow out of the vessel also removed. Lymph nodes have an important SELECTING A VEIN function for immunity. Upon removal of the breast and Use the tip of the index finger to palpate the vein the lymph attached to the lymph nodes, there is o Size lymphostasis. It makes that side of the body more o Depth probe to infection and swelling. o Direction Site of breast removal Select a vein o Lymphostasis o Easily palpated ▪ Infection o Large enough to support good blood flow ▪ Swelling o Well-anchored – we select the median-cubital o Lymphocytosis – Lymphocytes come from the vein lymph nodes and removal of these lymph nodes CLEANING THE SITE may produce a false elevation of the lymphocyte. Clean antiseptic ▪ WHAT DO WE DO? We can collect blood o MC: isopropyl alcohol (70%) from the contralateral or their opposite arm. “In to out” (concentric circles) (+) DOUBLE MASTECTOMY – both breasts and Use sufficient pressure to remove surface dirt. lymph nodes have been removed. We can collect Air dry the site alternatively from their hand vein or do a capillary Do not wipe or fan the site puncture. NEEDLE SIZE o Capillary puncture Correct guage: dec hemolysis OBESE o HOW TO SPOT A HEMOLYZED BLOOD The veins of these patients may be buried under fats SAMPLE? Hemolyzed samples would have red making them less visible and quite difficult to palpate. plasma due to hemoglobin that has been Need special collection released from the RBCs. o Veins not readily visible Larger the bore of the needle: < chance of hemolysis o Difficult to palpate o Smaller bores are usually used for smaller veins. Remedy for locating the vein Larger bores are used for larger veins. o Use of a blood pressure cuff (not higher than 40 NOTE: Needle gauge will vary based on patient size mmHg), not be left on the arm for >1 minute to o E.g., a smaller gauge needle is used for pediatric avoid hemoconcentration) patients ▪ Alternatively, we can use vein scanners to NEEDLE INSERTION visualize the veins only if available. Grasp the patient’s arm/hand with the thumb on top Phlebotomist should not probe blindly and fingers wrapped to the back. o Nerve damage or hematoma Pull the skin taut below the site with the thumb, TIPS IN VENIPUNCTURE anchoring. Tight enough to slow venous flow without affecting Smooth motion, quickly insert the needle, bevel up. arterial flow o With the hole facing upwards Allow more blood to flow into than out of the area FILLING OF TUBES Never be left longer than one minute APPLY FOR THE USE OF EVACUATED TUBE Not be applied on the arm of a recent site of a SYSTEM mastectomy Insert the collection tube o Applied on the opposite arm Maintain the tube in a downward position NOTE Fill the tube until the vacuum is exhausted o 3-4 inches above site o Tubes have a pre-determined vacuum that will fill o < 1 minute only the tube with blood until a certain volume. So, it RMT - 2024 11 HEMATOLOGY BSMLS 3F MLS 413 | LECTURE | PRELIMS 2022 - 2023 BLOOD COLLECTION usually stops filling up once the vacuum is used As soon as the needle is removed from the patient, up. the safety device on the needle must be activated Remove the tube from the holder by applying Immediately discard all needles in sharps container pressure NEEDLE STICK INJURY o by slightly twisting the tube while holding the MC injury in the lab needle in place as to not dislodge it from the vein. o Safety device not activated If tube (+) additive, invert it gently 8-10 times after o Improper disposal removal to mix the blood and additive. o Manipulating the needle in the patient INVERTING THE TUBES o Patient moves during the procedure. When: immediately after SAMPLE LABELING drawing specimen Never: Why: (+) additives o Never pre-label the tubes prior to blood collection How: holding tube upright, o Never leave a patient room before labeling the gently invert 180 degrees tubes and back Always Consequences if not mixed o Always verify: match the information (order slip properly and patient ID band) o (+) clot, re-draw o Label tubes after they are drawn in the presence o Vigorous, (+) hemolysis of the patient to reduce the risk of specimen When using a winged blood misidentification. collection set for venipuncture and a coagulation Note: fasting test must be labelled with the time of (citrate) tube is the first specimen tube to be drawn, a collection discard tube should be drawn first. The discard tube COMPLICATIONS OF VENIPUNCTURE must be used to fill the blood collection set tubing’s Some complications that can be encountered in “dead space” with blood, but the discard tube does venipuncture include hematoma or ecchymosis which not need to be completely filled. This important step may be prevented by avoiding blind shooting or will ensure proper blood-to-additive ratio. The discard probing tube should be a nonadditive or coagulation tube Dizziness which may lead to fainting or syncope may POST COLLECTION CARE be seen in patients especially those who have a fear Clean cotton/gauze of needles or trypanophobia Do not remove cotton too soon, disrupt forming clot Potential nerve damage can be avoided by avoiding Cover it with bandage blind shooting or probing Once the needle is removed, a clean cotton or gauze Hemolyzed blood samples can be prevented by using pad should be placed over the side and gentle the appropriately sized needle gauge and by inverting pressure should be applied for about two to five the tubes gently minutes to ensure that bleeding has stopped. Patients who have nausea or vomiting should be Do not remove the gauze or cotton too soon to avoid assisted disrupting the formed cloth. Secure the cotton with 1. Ecchymosis (bruise bandage or plaster 2. Hematoma DISPOSAL 3. Fainting (syncope) Never cut, bend, break, or recap needles to avoid 4. Hemoconcentration needle stick injury. 5. Hemolysis o This important because through needle stick 6. Petechiae injuries there is access to our blood and therefore 7. Allergies a potential risk of acquiring blood-borne diseases 8. Nerve damage like hepatitis B and HIV 9. Seizures o In a broad setting, some needles may have an 10. Vomiting attached safety device or a cap which must be CHALLENGES IN VENIPUNCTURE used upon removal of the needle from the skin. FAILURE TO DRAW BLOOD Vein is missed RMT - 2024 12 HEMATOLOGY BSMLS 3F MLS 413 | LECTURE | PRELIMS 2022 - 2023 BLOOD COLLECTION o You would know if the vein is hit appropriately For children or adults, we take we take blood from the when there is enough backflow into the needle's palmar surface of the distal portion of the third or hub fourth finger Insufficient vacuum in evacuated tube Remember to puncture perpendicular to the o It is important to check for the tube’s expiration fingerprint lines as to avoid spillage of the blood date to ensure its quality sideways Can also be caused by patient refusal. Therefore, it is EQUIPMENT USED IN CAPILLARY PUNCTURE important to establish good rapport with the patient 1. Safety lancet prior to blood collection 2. Pen or automatic lancet There may be cases wherein the patient is not found. Has a predetermined depth Therefore, there should be proper patient 3. Feather lancet identification by asking for the patient's name and 4. Microcontainer TUBE other pertinent information. TIPS IN CAPUILLARY PUNCTURE: FINGER PRICK ADEQUACY OF BLOOD VOLUME Finger or heel must be securely immobilized Blood volume is a critical factor in pediatric Heel punctures on infants should not be made more phlebotomy than 2mm deep o This is because even a small amount of blood may o Premature infants: 35 fL = WBC in the WBC/Hb chamber ○ Largest cell Lymphocytes = 35-90 fL ○ Mononuclear Mononuclears = 90-160 fL Between the T1 and T2 Neutrophils = 160-450 fL Eosinophilia Increase number of eosinophils ○ Bilobed Between the T1 and T2 It is very difficult to differentiate monocytosis Plotting the WBC size against their number. and eosinophilia in a WBC Histogram because: There are 3 analysis groups: ○ They are located/detected in the same ○ Lymphocytes region ○ Granulocytes ○ Perform peripheral blood smear to ○ Mononuclear cells differentiate the two Possibly that presence of malarial parasites in the RBC of a patient can cause an increase in the mixed cell population in the WBC histogram. The RBC that is infected by the malarial parasite HEMA - LEC COLO, REGULACION, YPIL|BSMLS 3I 4  HEMATOLOGY - LECTURE LESSON#2: HISTOGRAM MIDTERMS | A.Y. 2023 - 2024 | MA’AM VANESSA CALIMBO cannot be lysed by the stromatolyser, thus, they Poikilocytosis are able to enter the WBC counting block and ○ Variation in RBC shapes cause an increase in the mixed cell population. Presence of malarial parasites = unusual and predominant peak in the mixed cell population RED CELL HISTOGRAM It represents the relation between red cells size and the number ○ Routinely available on all automated cell RDW increased in counters ○ IDA Importance: ○ Megaloblastic anemia ○ Monitoring and interpreting abnormal ○ Hemoglobinopathies morphological changes in the RBCs, particularly dimorphic red cell RED CELL HISTOGRAM cont.. population which suggests RBC Platelets have volume between 8-12 fL and abnormalities. counted between 2-25 fL 2 parameters can be obtained: RBCs have volume 80-100 fL and are counted ○ Mean Cell Volume between 25-250 fL ○ Red Cell Distribution Width ○ Platelets and RBCs are in one histogram because they are counted in the same block If RBCs are larger than normal —> shift to right If RBCs are smaller than normal —> shift to left If the curve is bimodal —> 2 population of RBCs MEAN CELL VOLUME Broken line - fixed platelet discriminator and Calculated using the entire RBCs histogram is found lying at 12 fL Measures the average size of RBC ○ Normally, the RBC curve form a MCV decreased: gaussian curve (bell-shaped) ○ IDA ○ Thalasemia ○ Anemia of chronic diseases MCV increased: ○ Megaloblastic anemia ○ Acquired aplastic anemia RED CELL DISTRIBUTION WIDTH Coefficient of variation of RBCs volume distribution The discriminator is called RL. Measures the variation of RBC sizes ○ R for red blood cells ○ If RDW is increased, RBC population is ○ L for lower discriminator. more varied RL separates platelets from the RBC. Anisocytosis RU - Red Blood Cell Upper Discriminator ○ Variation of RBC sizes HEMA - LEC COLO, REGULACION, YPIL|BSMLS 3I 5  HEMATOLOGY - LECTURE LESSON#2: HISTOGRAM MIDTERMS | A.Y. 2023 - 2024 | MA’AM VANESSA CALIMBO ○ The y-axis of the histogram contains the Left picture - hypochromic (faint-colored), relative number of cells being plotted. central pallor is big The distributional shape of the histogram can be ○ Majority of the cell population are classified as: microcytic and causes a shift to the left ○ Single population in the RBC histogram ○ Dimorphic ○ Multiple RBCs SHIFT TO THE RIGHT Dimorphic RBCs ○ 2 or more types of RBC population seen in the peripheral blood sample Ex. (1) Microcytic - hypochromic RBCs, (2) Macrocytic - hyperchromic RBCs, (3) Normocytic - normochromic Macrocytosis RBCs ○ Macrocyte population is predominant ○ Expect a dimorphic RBC population after ○ There are plenty of enlarged RBCs in the iron treatment when the patient has IDA peripheral blood smear of the patient ○ Symmetrical or bimodal but skewed If there is a skewing of the curve to the right, it is histogram suggestive to predominant macrocytic RBCs, especially macrocyte population Single population ○ Megaloblastic Anemia ○ Normal gaussian curve but may be ○ Acquired Aplastic Anemia widened without skewing towards the left or right (shifting) ○ Skewing towards the left or right Possible RBC anomalies ○ The centeredness and the width of the histogram define the extent of the RBC variability SHIFT TO THE LEFT Left picture - predominant of macrocytes, macrocytes have a small central pallor (Observe Lymphocyte at the center) Right picture - predominant of macrocytic Red Blood Cells (Observe Lymphocytes at the center) DIMORPHIC POPULATION Microcytosis - skewed to the left ○ The lesser the fL, the smaller the RBC size If we see this, it is suggestive of Dimorphic Population. There is a predominance of 2 types of population or even 3. Right picture - normal-sized and shaped RBCs, It could be a mix of: central pallor is small ○ Microcytic Hypochromic Cells ○ Macrocytic Hyperchromic Cells HEMA - LEC COLO, REGULACION, YPIL|BSMLS 3I 6  HEMATOLOGY - LECTURE LESSON#2: HISTOGRAM MIDTERMS | A.Y. 2023 - 2024 | MA’AM VANESSA CALIMBO ○ Normocytic Normochromic Cells ○ Microcytic Hyperchromic Cells Photo A - Peripheral Blood Smear with a dimorphic population of RBC which results to be bimodal in the graph. We can see in the picture the ○ Normocytic RBC ○ Normocytic Normochromic ○ Microcytic Hypochromic ○ Macrocytes ○ Tear drop cells and; ○ Schistocytes Eg. If you're able to see these in the smear (Microcytic and Hypochromic RBCs + Normocytic Normochromic) ○ The patient could have an Iron Deficiency Anemia and is undergoing Iron treatment. That is the reason for having a mixed population of RBCs in the peripheral blood sample. [END] HEMA - LEC COLO, REGULACION, YPIL|BSMLS 3I 7  Topic 2 | [MLS 417-LAB] Hematology 2 - Lecture P2: Introduction to RBC Abnormalities Professor: Lee-An Anayon, RMT Date: February 11, 2024 IMPORTANT TERMINOLOGIES ERYTHROCYTOSIS AND POLYCYTHEMIA (Increased RBC) ABSOLUTE VS. RELATIVE (↓) Decreased Hemoglobin ABSOLUTE ○ Due to the dilution effect of the increased In absolute anemia or polycythemia there is a TRUE RBCs DECREASE or INCREASE in the Red Cell Mass (↑) Increased Hematocrit (RCM), respectively. (↑) Increased Number of Red Cells ○ If True anemia – there is really a problem in the RBC; Lysis occurs; rapid destruction of RBC ○ RCM refers to the volume of the plasma RELATIVE Changes in plasma volume causing changes in cellular components to cellular components ○ There is a shift of fluid from extracellular to intracellular – The blood will be diluted ○ Decreased RBC count due to the dilution phenomenon that happened during the changes in plasma volume ○ It does not mean that the patient has true anemia ERYTHROCYTE ABNORMALITIES Relative anemia – pregnancy and diseases A. Decreased Concentration associated with hyperproteinemia Ineffective/Insufficient erythrocyte production ○ Relative anemia occurs in pregnancy due to ○ Hypoproliferative disorders plasma volume expansion, making RBC Precursor cells inside the bone concentration appear low without a true RBC marrow has low count; cannot deficiency. produce now enough RBC ○ Diseases with hyperproteinemia involve ○ Maturation disorders excessive blood protein levels, potentially Bone marrow is incapable of affecting blood viscosity and composition, producing mature RBC but don't necessarily decrease RBC count. Our RBCs are only released in the Relative erythrocytosis – dehydration peripheral blood only when it ○ In this state (dehydrated state), the matures – except in cases of blood concentration of RBCs appears elevated loss where reticulocytes are because there's less plasma to dilute them, released early even though the actual number of RBCs may Increased RBC destruction/ Loss remain normal. Nothing wrong with the bone marrow but RBC destruction is rapid such in cases of: ○ Hemolytic disorders ○ Blood loss – menstruation That is why in cases when we will have a reticulocyte count in the laboratory our sample should come from our classmates who is in her period ANEMIA, ERYTHROCYTOSIS AND POLYCYTHEMIA ANEMIA (Decreased RBC) (↓) Decreased Hemoglobin ○ Due to increased destruction of RBC the Hgb are released (↓) Decreased Number of Red Cells ○ Still due to increased destruction of RBC Why is it important to diagnose the cause of anemia? Why must it be prompt? Because sometimes the presence of anemia is not alone in itself, it may have underlying diseases, that’s why we have anemia; It must be prompt, for the doctors to give the final Anemia can be categorized into 4 as mentioned in the table above diagnosis and for the immediate resolution of the disease @mlstranses | 1  B. Increased Concentration SECONDARY (High altitude) When you live in a high altitude area your hematocrit is increased to compensate the oxygen that the body needs RELATIVE When you smoke tobacco you inhale carbon dioxide and not MODERATE ANEMIA oxygen, so your body needs to compensate to balance the Hemoglobin concentration of 7-10 g/dL carbon dioxide and the oxygen May cause pallor of conjunctiva and nail beds ANEMIA SEVERE ANEMIA Anemia comes from the greek word ANAIMA which means Hemoglobin concentration of 8 um in Megaloblastic anemia diameter), MCV > 100 fL Myelodysplastic syndromes Chronic liver disease Bone marrow failure Reticulocytosis Oval macrocyte Large oval RBC Megaloblastic anemia Microcyte Small RBC (