Laboratory-General Specimen Collection and Handling Guidelines
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Handout Page 1 of 8
MLT 112: Principles of Phlebotomy Learning Unit 3: Handout Specimen Collection – Venipuncture Procedure I. Collecting and Processing of Specimens A. Blood 1. Venipuncture Procedure (arm + dorsal/hand) – vacutainer, syringe, butterfly a. Approaching the Patient Correct patient identification Wash hands Have patient recite his/her name Wrist identification – mandatory must match requisition check ankle on babies and peds Out-patients – ask patient to spell name In-patients – see wrist ID Unconscious patient – see wrist ID Unidentified patient – emergency – use temporary I.D. band non-emergency – wait for I.D. Explanation and Reassurance – Inspire confidence Conversation Ensure that patient has complied with test requirements, such as fasting (only water), NPO (nothing per oral), etc. Check for any allergies, such as to latex, adhesive bandages, etc. b. Positions Positioning the patient Vein accessibility Sitting vs. lying down Phlebotomist position – always in front in case of fainting. c. Applying the Tourniquet See previous lecture Page 1 of 8 MLT 112: Principles of Phlebotomy Learning Unit 3: Handout d. Veins Used - Antecubital Fossa Cephalic Median Cephalic Median Basilic Median Cubital Vein – vein of choice, anchored best Other Structures – avoid Brachial artery – apply pressure 5 minutes Cutaneous nerve – very painful Tendon for the biceps muscle – always draw below crease e. Other Vein Sites Wrist (never palm side) Hand Ankle Foot f. Preparing Equipment Syringes Assembly – always work plunger before procedure. Plunger position – must -
Histopathology and Laboratory Features of Sexually Transmitted Diseases
Histopath & Labs for STIs Endo, Energy and Repro 2017-2018 HISTOPATHOLOGY AND LABORATORY FEATURES OF SEXUALLY TRANSMITTED DISEASES Dominck Cavuoti, D.O. Phone: 469-419-3412 Email: [email protected] LEARNING OBJECTIVES: • Identify the etiologic agents causing pelvic inflammatory disease and the pathologic changes they produce. • Discuss the characteristic clinical and pathologic findings caused by herpes simplex virus (HSV) infections: a. fever blisters b. genital herpes simplex virus infection c. disseminated neonatal HSV • Describe the pathologic changes produced by Treponema pallidum. • Describe the clinical features and pathologic changes produced by Chlamydia trachomatisand Neisseria gonorrhoeae • Describe the clinical and laboratory features of vaginal infections including: Trichomonas, Candida, and bacterial vaginosis. • Describe the clinical and laboratory features of ectoparasite infections PURPOSE OF THE LECTURE: 1. To describe the various agents of sexually transmitted diseases and their disease manifestations 2. To describe the pathologic features associated with STDs 3. To introduce some of the laboratory aspects of STDs TERMS INTRODUCED IN LECTURE: Condyloma lata Disseminated gonococcal infection Gummatous syphilis Lymphogranuloma venereum Pelvic inflammatory disease Rapid Plasma Reagin (RPR) Salpingitis Syphilis/endarteritis obliterans Venereal Disease Research Laboratory (VDRL) Treponema pallidum particle agglutination (TPPA) Histopath & Labs for STIs Endo, Energy and Repro 2017-2018 MAJOR CONCEPTS EMPHASIZED IN LECTURE I. Syphilis (Will be covered by Dr. Norgard in later lecture). II. Gonorrhea A. Causative agent: Neisseria gonorrhoeae, a Gram negative diplococcus. Humans are the only natural reservoir. Infection is acquired via direct contact with the mucosa of an infected person. The incubation period averages 2-5 days with a range of 1-14 days. -
Vacutainer Tubes
Vacutainer Tube Guide - Order of Draw for routine volumes See separate guide for micro-tainers Number of Tube Inversions at Label Blood Collection Volume Tubes/Bottles Abbrev. Additive (Do Not Shake) General Laboratory Use Draw 1 Aerobic and 1 5 Blood Culture / requires Adapter 20 mL Anaerobic Culture Note: A separate venipuncture for trace BLCLT Bottles metal analysis is required if blood cultures are ordered at the same time. NAYYP Clot activator 8 Trace Element serum tube for Copper 6 mL Royal Trace Element whole blood for Aluminum, NAVYE 2 Call 2-5002 Blue K EDTA Lead, Zinc, Selenium, Manganese, Mercury, 8 for tubes Cadmium, Arsenic Discard tube or secondary sterile specimen 6 mL Clear None 0 tube 3 mL 2.7 mL Light (Blue) BLUE Sodium Citrate (3.2%) Coagulation Testing on Plasma Blue 3-4 1.8 mL (Blue/clear) Serum determinations where a gel-barrier is 6 mL Red RED Clot Activator 5 contraindicated; Therapeutic Drug 4 mL Monitoring (TDM) Clot Activator and gel SST - Serum determinations in Chemistry, Gold SST 5 5 mL for serum separation Immunology, Serology; HLA Ab screen, DSA Light Lithium heparin and PST PST - Plasma determinations in Chemistry 4.5 mL Green gel - plasma separation 8 GLI Lithium heparin 8 Whole Blood Plasma: Troponin, Lactic Acid, Ion. Ca+ 6 mL Dark 4 mL Green GNA Sodium heparin 8 Whole Blood Plasma: Tissue typing, plasma hemoglobin, plasma catecholamine, chromosome analysis Pink PINK Dry K2EDTA 8 Blood Bank / Transfusion; HLA Typing 6 mL Tan TAN K2EDTA 8 Lead Testing only 3 mL LAV 4 Whole Blood Hematology; some Flow, 4 mL Lavender Dry K2EDTA 8 Molecular, and Genetic tests; HIV Viral 2 mL LAV6 Load Whole Blood tube with Glycolytic inhibitor Potassium Oxalate / for GTT, Gray 6 mL GRAY Sodium fluoride 8 Post-Prandial Glucose, D-Xylose, and Volatiles panel ACDA ACD Whole Blood tube for Flow Yellow (Acid Citrate Dextrose 8 immunophenotyping; HLA cross matching; 8.5 mL ACDB Solution A) genetic and other referred tests ALWAYS Check Expiration Dates before use! If unsure of use, refer to the on-line “Lab User’s Guide” or call the lab at 25002 . -
Coagulation Testing: High Hematocrit-Anticoagulant Adjustment
PREANALYTIC PULSE Coagulation Testing: High Hematocrit-Anticoagulant Adjustment In 1980, CLSI released H21-A4 guidelines for coagulation testing, which included the recommendation to adjust/correct the amount of citrate in blue-top evacuated blood collection tubes for patients presenting hematocrits greater than 55%. In addition to the CLSI guidelines, the CAP hematology checklist HEM.22830 includes the question, “Are there documented guidelines for the detection and special handling of specimens with elevated hematocrits?” Principle The adjustment is to assure the plasma:anticoagulant ratio (not the blood:anticoagulant ratio) stays consistent. A patient with high hematocrit, greater than 55%, will result in less plasma after centrifugation. The plasma fraction will contain an increased concentration of sodium citrate anticoagulant. The increased concentration may result in falsely prolonged test results for Prothrombin Time (PT) and Activated Partial Thromboplastin Time (aPTT). Formula 1. To calculate the corrected 3.2% sodium citrated whole blood for hematocrit > 55%, adjust the citrate to the proper volume with the following formula: -3 C = (1.85 x 10 )(100-HCT)(Vblood) Where: C = volume of sodium citrate required for that volume of blood HCT = patient’s hematocrit V = volume of blood required in the blood collection tube (example if a 3mL tube is used the blood draw volume is 2.7mL) 1.85 x 10-3 is a constant (considering the citrate volume, blood volume, and citrate concentration). 2. Example: Patient has a Hct of 60% and the patient’s blood will be drawn into a 3mL VACUETTE® sodium citrate (blue-top) blood collection tube. Adjustment of the sodium citrate volume is calculated as: C = (1.85 x 10-3)(100-60)(2.7mL) C = 0.20mL (rounded up from 01.998mL) Remove: 0.30 - 0.20 = 0.10mL of sodium citrate to be removed (0.30 is the difference between the total tube volume of 3.0mL and the blood drawn into the tube of 2.7mL) Method The instructions to prepare an adjusted sodium citrate tube are to be documented in the Laboratory Standard Operating Procedures. -
Francisella Tularensis 6/06 Tularemia Is a Commonly Acquired Laboratory Colony Morphology Infection; All Work on Suspect F
Francisella tularensis 6/06 Tularemia is a commonly acquired laboratory Colony Morphology infection; all work on suspect F. tularensis cultures .Aerobic, fastidious, requires cysteine for growth should be performed at minimum under BSL2 .Grows poorly on Blood Agar (BA) conditions with BSL3 practices. .Chocolate Agar (CA): tiny, grey-white, opaque A colonies, 1-2 mm ≥48hr B .Cysteine Heart Agar (CHA): greenish-blue colonies, 2-4 mm ≥48h .Colonies are butyrous and smooth Gram Stain .Tiny, 0.2–0.7 μm pleomorphic, poorly stained gram-negative coccobacilli .Mostly single cells Growth on BA (A) 48 h, (B) 72 h Biochemical/Test Reactions .Oxidase: Negative A B .Catalase: Weak positive .Urease: Negative Additional Information .Can be misidentified as: Haemophilus influenzae, Actinobacillus spp. by automated ID systems .Infective Dose: 10 colony forming units Biosafety Level 3 agent (once Francisella tularensis is . Growth on CA (A) 48 h, (B) 72 h suspected, work should only be done in a certified Class II Biosafety Cabinet) .Transmission: Inhalation, insect bite, contact with tissues or bodily fluids of infected animals .Contagious: No Acceptable Specimen Types .Tissue biopsy .Whole blood: 5-10 ml blood in EDTA, and/or Inoculated blood culture bottle Swab of lesion in transport media . Gram stain Sentinel Laboratory Rule-Out of Francisella tularensis Oxidase Little to no growth on BA >48 h Small, grey-white opaque colonies on CA after ≥48 h at 35/37ºC Positive Weak Negative Positive Catalase Tiny, pleomorphic, faintly stained, gram-negative coccobacilli (red, round, and random) Perform all additional work in a certified Class II Positive Biosafety Cabinet Weak Negative Positive *Oxidase: Negative Urease *Catalase: Weak positive *Urease: Negative *Oxidase, Catalase, and Urease: Appearances of test results are not agent-specific. -
Biofire Blood Culture Identification System (BCID) Fact Sheet
BioFire Blood Culture Identification System (BCID) Fact Sheet What is BioFire BioFire BCID is a multiplex polymerase chain reaction (PCR) test designed to BCID? identify 24 different microorganism targets and three antibiotic resistance genes from positive blood culture bottles. What is the purpose The purpose of BCID is to rapidly identify common microorganisms and of BCID? antibiotic resistance genes from positive blood cultures so that antimicrobial therapy can be quickly optimized by the physician and the antibiotic stewardship pharmacist. It is anticipated that this will result in improved patient outcomes, decreased length of stay, improved antibiotic stewardship, and decreased costs. When will BCID be BCID is performed on all initially positive blood cultures after the gram stain is routinely performed and reported. performed? When will BCID not For blood cultures on the same patient that subsequently become positive with be routinely a microorganism showing the same morphology as the initial positive blood performed? culture, BCID will not be performed. BCID will not be performed on positive blood cultures with gram positive bacilli unless Listeria is suspected. BCID will not be performed on blood culture bottles > 8 hours after becoming positive. BCID will not be performed between 10PM-7AM on weekdays and 2PM-7AM on weekends. BCID will not be performed for clinics that have specifically opted out of testing. How soon will BCID After the blood culture becomes positive and the gram stain is performed and results be available? reported, the bottle will be sent to the core Microbiology lab by routine courier. BCID testing will then be performed. It is anticipated that total turnaround time will generally be 2-3 hours after the gram stain is reported. -
Interpreting and Acting on Positive Blood Cultures Trevor Van Schooneveld, MD 1/18/18 Objectives
Stewardship Interventions: Interpreting and Acting on Positive Blood Cultures Trevor Van Schooneveld, MD 1/18/18 Objectives • Interpret the results of blood cultures including gram stains and rapid pathogen diagnostic tests • Make recommendations regarding antimicrobial therapy based on interpretation of blood culture data Early Initiation of Active Therapy is Essential Predicted hospital mortality and 95% CIs for time to first antibiotic administration Surviving Sepsis Guidelines (N=28,150 severe sepsis, septic shock patients) • Administer IV antimicrobials within one hour of presentation (strong) • Initiate empiric, broad-spectrum therapy with one or more agents to cover all likely pathogens (strong) Ferrer R, et al. Crit Care Med. 2014;42:1749-55. Rhodes A, et al. Crit Care Med. 2017;45:486-552. De-escalation Also is Important Surviving Sepsis Guidelines • Narrow empiric antibiotics once pathogen identified and/or clinical improvement De-escalation Benefit • De-escalation in severe sepsis, septic shock (N=712) • Mortality OR 0.54 (95% CI 0.33-0.89, P=.016) • De-escalation in community-onset gram-negative bacteremia (N=189) • Mortality OR 0.37 (0.14-0.96, P=.04) Garnarcho-Montero J, et al. Intensive Care Med. 2014;40:32-40. Lee C, et al. Diag Micro Infect Dis. 2015;82:158-64. Issues with Treatment of Sepsis/Bacteremia Under-treatment • May die (mortality) • May not get better as quickly (LOS, cost) • May develop complications (LOS, cost) Overtreatment • May develop toxicities (cost, LOS) • May develop C. difficile (cost, LOS, readmission) -
Role of Real-Time / Accurate Diagnostic Testing in Patient Blood Management Programs
ROLE OF REAL-TIME / ACCURATE DIAGNOSTIC TESTING IN PATIENT BLOOD MANAGEMENT PROGRAMS POINT-OF-CARE TESTING “Diagnostic testing is an essential component of Patient Blood Management. The accurate assessment of the true causes of bleeding dysfunction facilitates the employment of evidence based, goal directed therapy to rapidly prevent and treat excess blood loss. “ Sherri Ozawa RN, Clinical Director, Institute of Patient Blood Management, Englewood Hospital Medical Center, Englewood, New Jersey. PATIENT BLOOD MANAGEMENT Interdisciplinary Managing Blood Conservation Anemia Modalities “The timely application of evidence based medical and surgical concepts designed to manage IMPROVED anemia, optimize hemostasis, and PATIENT minimize blood loss in order to OUTCOMES improve patient outcomes.” SABM© 2018 - Society for the Advancement of Blood Management (SABM.org) Optimizing Patient-Centered Coagulation Decision Making Point-of-care testing forms an integral part of PBM, enabling accurate and real-time diagnosis of anemia and bleeding and facilitating precise and targeted hemostatic intervention. 1 Conventional coagulation tests are unable to provide needed information on actual bleeding risk/defects 1, thus delaying appropriate and targeted treatment. Slow turnaround times for laboratory-based conventional coagulation tests are one of the major drivers of empirical treatment regimens, which inevitably result in some patients receiving unnecessary, inappropriate and avoidable transfusions, with associated increased morbidity and mortality, and -
BLOOD CULTURE MEDIA Principle: Specimen: Reagent Preparation Storage: Procedure
BLOOD CULTURE MEDIA For In-Vitro and professional use only Store at (2° to 8°C) Blood cultures are used to detect the presence of bacteria or fungi in the blood, to identify the type present, and to guide treatment. Testing is used to identify a blood infection (septicemia) that can lead to sepsis, a serious and life-threatening complication. Individuals with a suspected blood infection are often treated in intensive care units, so testing is often done in a hospital setting. A bacterial infection in the blood called bacteremia. It can be serious because the blood can spread the bacteria to any part of the body. Blood infections most often occur with other serious infections such as those affecting the lungs, kidneys, bowel, gallbladder, or heart valves. Blood infections may also develop when the immune system is weak in infants and older adults, from disease (such as cancer or AIDS) or from medicines (such as corticosteroids or chemotherapy) that change the ability of your body to fight infections (immunity). Principle: The vials containing 25 ml or 50 ml of brain heart infusion, yeast extract, SPS and other stabilizers. The Media is used for yeast, aerobic and anaerobic organisms in blood. The principle of the this test is that each type of organisms need a certain time to grow and multiply. Specimen: Blood. Reagent preparation The vials are ready to use. Storage: Store reagent from (2 - 8 oC). Procedure: 1. Bottles of Brain Heart Infusion which are not used the same day as sterilized should be placed in a boiling water bath for several minutes to remove absorbed oxygen , and cooled rapidly without shaking , just before use. -
Global Assays of Hemostasis in the Diagnostics of Hypercoagulation and Evaluation of Thrombosis Risk Elena N Lipets1 and Fazoil I Ataullakhanov1,2,3,4,5,6*
Lipets and Ataullakhanov Thrombosis Journal (2015) 13:4 DOI 10.1186/s12959-015-0038-0 REVIEW Open Access Global assays of hemostasis in the diagnostics of hypercoagulation and evaluation of thrombosis risk Elena N Lipets1 and Fazoil I Ataullakhanov1,2,3,4,5,6* Abstract Thrombosis is a deadly malfunctioning of the hemostatic system occurring in numerous conditions and states, from surgery and pregnancy to cancer, sepsis and infarction. Despite availability of antithrombotic agents and vast clinical experience justifying their use, thrombosis is still responsible for a lion’s share of mortality and morbidity in the modern world. One of the key reasons behind this is notorious insensitivity of traditional coagulation assays to hypercoagulation and their inability to evaluate thrombotic risks; specific molecular markers are more successful but suffer from numerous disadvantages. A possible solution is proposed by use of global, or integral, assays that aim to mimic and reflect the major physiological aspects of hemostasis process in vitro. Here we review the existing evidence regarding the ability of both established and novel global assays (thrombin generation, thrombelastography, thrombodynamics, flow perfusion chambers) to evaluate thrombotic risk in specific disorders. The biochemical nature of this risk and its detectability by analysis of blood state in principle are also discussed. We conclude that existing global assays have a potential to be an important tool of hypercoagulation diagnostics. However, their lack of standardization currently impedes their application: different assays and different modifications of each assay vary in their sensitivity and specificity for each specific pathology. In addition, it remains to be seen how their sensitivity to hypercoagulation (even when they can reliably detect groups with different risk of thrombosis) can be used for clinical decisions: the risk difference between such groups is statistically significant, but not large. -
Gram Stain Workshop for the Laboratory Generalist
Gram Stain Workshop for the Laboratory Generalist Karen Stiles, SM(ASCP)CM State Training Coordinator Assistant Chemical Terrorism Coordinator Nebraska Public Health Laboratory 402-559-3590 [email protected] 1 GRAM STAIN OBJECTIVES: Upon completion, the participant will be able to: 1. Explain the principle of the Gram stain procedure, including what elements can affect staining results 2. Correlate the most common pathogens with positive Gram stains from blood cultures and direct specimen sterile body fluid smears 3. Perform and interpret Grams stains 2 Purpose of Gram Stain Classify bacteria based on form, size, cellular morphology, Gram reaction Assess quality of specimen Identify specific infectious agent from morphology and Gram reaction Correlation with culture growth Correlation with culture-independent methodologist Guide presumptive antibiotic therapy 3 Principle of Gram Stain Cell wall composition Gram positive – think peptidoglycan layer with teichoic acid Gram negative – high in lipid content Basic premise Crystal Violet – all cells take up primary stain Gram’s iodine – mordant to form complex Decolorizer – mixture of acetone and alcohol Dehydrate lipids in Gram negative cell walls, wash out complex Gram positive cells resistant, retain stain complex Safranin - counterstain 4 Gram negative cells take up counterstain Preparation of Samples Specimen Type Preparation CSF/sterile body fluids Cyto/Centrifuge Blood Culture Broth Drop to slide Tissue Touch prep Tissue homogenate Drop to slide Swabbed material Roll -
64Th Annual SSC Meeting, in Conjunction with the 2018 ISTH Congress in Dublin, Ireland Meeting Minutes Standing Committees Coagulation Standards Committee
64th Annual SSC meeting, in conjunction with the 2018 ISTH Congress in Dublin, Ireland Meeting Minutes Standing Committees Coagulation Standards Committee............................................................... 3 Subcommittees Animal, Cellular and Molecular Models ........................................................ 6 Biorheology .................................................................................................. 9 Control of Anticoagulation ............................................................................ 15 Disseminated Intravascular Coagulation ...................................................... 20 Factor VIII, Factor IX and Rare Coagulation Disorders ................................ 23 Factor XI and the Contact System ............................................................... 27 Factor XIII and Fibrinogen ............................................................................ 30 Fibrinolysis ................................................................................................... 34 Genomics in Thrombosis and Hemostasis ................................................... 38 Hemostasis and Malignancy ........................................................................ 47 Lupus Anticoagulant/Phospholipid Dependent Antibodies ........................... 51 Pediatric and Neonatal Hemostasis and Thrombosis ................................... 57 Perioperative and Critical Care Thrombosis and Hemostasis ...................... 66 Plasma Coagulation Inhibitors .....................................................................