1	CLSC320 Principles of Immunology Diagnostic Laboratory Immunology Program for Clinical Laboratory Science Unit - 09 Lab Safety & Test Quality Assurance
2	Unit – 09 Guidelines Reading assignment: Pages 192 - 211 of textbook Learning objectives: Those listed on page 193 of textbook Key terms: Those listed on pages 193 & 194 of textbook
3	Gov’t Agencies & Regulatory Function Gov=t Agency Regulatory Function OSHA OSHA = Occupational Safety & Health Administration CDC CDC = Centers for Disease Control & Prevention EPA EPA = Environmental Protection Agency HCFA HCFA = Health Care Financing Administration JCAHO JCAHO = Joint Commission on The Accreditation of Healthcare Organizations CAP CAP = College of American Pathologists guidelines for bloodborne pathogens safety standards for handling infectious agents management of biohazardous wastes sponsor of CLIA=88 overall quality of laboratory improvement of services & test results
4	Protective Measures for Workers Barrier protection: Preventive measures: UVinyl or latex gloves Ugowns or lab coats (impermeable) Uface shields or masks & eye protectors Uwash hands with soap & water after removing gloves Uwash with soap & water after exposure to hazardous materials Uno mouth pipeting Udecontaminate work areas with 1:10 dilution bleach Udispose of contaminated materials in appropriate biohazard labelled containers Uno eating or drinking in lab area Uwash hands & remove protective clothing before leaving lab
5	Universal Precautions Definition: The handling of all human blood or other potentially infectious materials as though they are contaminated with bloodborne pathogens and are potentially infectious. Exposure Control Plan Definition: Each institution is responsible for developing its own plan to assist emplyees in dealing with exposure to hazardous materials. Plan must include: UStandard Operating Procedures Manual (SOP) UChemical Hygiene Plan (CHP)) UEducation Program on Safety procedures & policies
6	Standard Operating Procedures Exposure Control Plan: continued Standard Operating Procedures (SOP) Manual: Ulisting of all lab tests and other procedures that present a potential exposure to infectious agent Uexplanation on ways to control the risk of exposure Uprovision for personnel education with a focus on preventing exposure to infectious agents Usignature of reviewer, date, and any updates to SOP
7	Chemical Hygiene Plan Exposure Control Plan: continued Chemical Hygiene Plan (CHP): Uwritten policies, procedures, and responsibilities to ensure that exposure to hazardous chemicals is prevented or minimized. Uyearly physical inventory of all hazardous chemicals UMaterial Safety Data Sheets (MSDS) for all hazardous chemicals must be available at all times econtain product information supplied by manufacturer giving hazardous status of chemical and steps to take if exposure occurs. Uall hazardous chemicals must be labelled with appropriate biohazard symbol
8	Universal Biohazard Symbol Exposure Control Plan: continued Universal Biohazard Symbol
9	Universal Biohazard Label Universal Biohazard Label Fire Hazard Health Hazard Reactivity Specific Hazard
10	Fire Hazard Label Universal Biohazard Label Fire Hazard flash points 4 = < 73^oF 3 = 73^oF - 100^oF 2 = 100^oF - 200^oF 1 = > 200^oF 0 = will not burn 4
11	Health Hazard Label Universal Biohazard Label Health Hazard 4 = Deadly 3 = Extreme danger 2 = Hazardous 1 = Slightly hazardous 0 = normal material 4
12	Specific Hazard Label Universal Biohazard Label OXY = oxidizer ACID = acid ALK = alkaline COR = corrosive Specific Hazard W = use no water = radiation
13	Reactivity Hazard Label Universal Biohazard Label 4 = may detonate Reactivity 3 = shock & heat may detonate 2 = violent chemical change 1 = unstable if heated 0 = stable 4
14	Hazardous Material Management Infectious waste: Uany waste that has been contaminated with blood or body fluids is considered a biohazard waste and is treated same as the blood or body fluids. Ubiohazardous waste must be incinerated or autoclaved Ua written contract is required between the hospital and disposal company defining how biohazardous waste will be handled and discarded. Chemical waste: Uany hazardous chemical waste must be handled, transported, and disposed of according to current EPA regulations.
15	Method Evaluation Assumptions regarding laboratory testing: Ulaboratory practitioner must demonstrate competency for procedure and instrumentation. Ureagents, controls, and calibrating materials used to manufacturer=s protocol. Ucommercially prepared materials are being used prior to the expiration period. Uthe method shows analytic linearity throughout the reporting range. Method evaluation should include: Utesting for Arandom error (RE)@ Utesting for Asystematic error (SE)@
16	Testing For Random Error Uerrors in testing that affect the performance or reliability of a procedure: Ucaused by variability in: etemperature ereagents and calibrators etechnical performance of testing personnel apipeting atiming amixing einstrument stability
17	Testing For Systematic Error Uerrors in testing that affect the performance or reliability of a procedure and may be constant or proportional: econstant systematic error (CE): eproportional systematic error (PE): aerror is consistently low or high by same amount and is independent of concentration. -interfering substances -instrument malfunction aerror is consistently low or high in proportion to concentration. -incorrect calibration
18	Testing For Total Error Uestimated by combining random and systematic error. Method Comparisons: U40 - 100 patient specimens are tested by running: eon a known accurate method and current method eon a current method and new method Uperform statistical analysis on data: et-test eCorrelation coefficient 0 = no correlation +1 = perfect correlation eLinear regression
19	Precautions & Specimen Handling Collection of specimens: Ucollected in appropriate containers Uhemolysis of specimen to be avoided Uproper specimen identification must include: ePatient name ePatient ID number number is unique to patient ePhlebotomist ID eDate & Time of collection eBlood Bank number Uproper specimen transport & storage to be followed Procedure for inactivating complement: Uheat at 56^oC for 30 minutes
20	Specimen Dilutions Preparation of diluted specimens or solutions: Udilution is an expression ot the concentration of a sample in a specific volume Udiluent is the solution used to dilute a sample
21	Solutions - 1 A.Basic ways of measuring concentration of solutions: 1.Weight per unit weight (w/w) 2.Weight per unit volume (w/v) 3.Volume per unit volume (v/v) B.Calculations of solutions: 1.Weight per unit weight (w/w) Make 500 gm of a 10% NaCl aqueous solution 500 gm x 0.10 = 50 gm then add the 50 gm NaCl to 450 gm H₂O
22	Solutions - 2 B.Calculations of solutions cont=d: 2.Weight per unit volume (w/v) Make 1000 ml of a 0.85% NaCl aqueous solution 0.85 gm x x gm 100 ml 1000 ml 100x = 850 x = 8.5 gm so you would put 8.5 gm NaCl in flask and add H₂O to 1000 mark.
23	Solutions - 3 B.Calculations of solutions cont=d: 3.Volume per unit volume (v/v) Make 500 ml of a 5% HCl using stock 10% HCl solution V₁ x C₁ = V₂ x C₂ 500 x 5 = X x 10 x = 250 ml so you would put 250 ml of 10% HCl in flask and add H₂O to 500 mark.
24	Dilutions - 1 A.Definition Procedures in which an amount of one substance is added to another to reduce the concentration of one of the substances. A dilution is an expression of concentration, not volume. It indicates the relative amount of the substances in a solution. B.Rules In dilution statements, the smaller number is the number of parts of the substance that is being diluted; the larger number is the total number of parts in the final solution.
25	Dilutions - 2 C.Ways to express the addition of 1 ml of serum to 9 ml saline. Procedures in which an amount of one substance is added to another to reduce the concentration of one of the substances. 1 to 10 dilution of serum in saline 1 in 10 dilution of serum in saline 1 to 10 dilution of serum with saline 1/10 dilution of serum with saline 1:10 dilution of serum using saline 1 part serum and 9 parts saline *1 part serum to 9 parts saline
26	Dilutions - 3 D.Calculation examples: 1.Dilute 3 ml serum with 25 ml saline Dilution = volume of saline + volume of serum volume of serum Dilution = 25 + 3 3 Dilution = 9.33 or 1:9.33 or 1 part serum to 8.33 parts saline 2.Dilute 5 ml blood with 25 ml saline Dilution = 25 + 5 5 Dilution = 6 or 1:6 or 1 part blood to 5 parts saline
27	Serial Dilutions - 1 A.Definition A series of dilutions in which all dilutions after the first are the same. Example:a series of 5 test tubes are set up, each containing 2 ml of saline. Into the first tube 1 ml of serum is added. After mixing, 0.5 ml of tube 1 is transferred into tube 2. This is repeated for each remaining tube. First calculate dilution in tube 1 Dilution = volume of saline + volume of serum volume of serum Dilution = 2 + 1 = 1:3 or 1/3 1
28	Serial Dilutions - 2 Next calculate dilution in tube 2 Dilution = 2 + 0.5 = 1:5 or 1/5 0.5 so to calculate the serial dilution you multiply the dilution factor of each tube: 1 x 1 x 1 x 1 x 1 = 1 3 5 5 5 5 1875
29	Serial Dilutions - 3 A.Definition A series of dilutions in which dilutions vary from tube to tube. Example:a series of 6 test tubes are set up, each containing 2 ml of saline. Into the first tube 1 ml of serum is added. After mixing, 0.5 ml of tube 1 is transferred into tube 2. This is repeated for tubes 3,4, and 5. Only 0.2 ml of tube 5 is transferred to tube 6 First calculate dilution in tube 1 Dilution = volume of saline + volume of serum volume of serum Dilution = 2 + 1 = 1:3 or 1/3 1
30	Serial Dilutions - 4 Next calculate dilution in tube 2 thru 5 Dilution = 2 + 0.5 = 1:5 or 1/5 0.5 Next calculate dilution in tube 6 Dilution = 2 + 0.2 = 1:11or 1/11 0.2 so to calculate the serial dilution you multiply the dilution factor of each tube: 1 x 1 x 1 x 1 x 1 x 1 = 1 3 5 5 5 5 11 20,625
31	Titer antibody titer is an expression that refers to the concentration of antigen-specific antibodies in a particular serum dilution. Uit is reported as the highest (last) dilution in which the antigen-antibody reaction is visible (1+) using a serial dilution technique. Ublood samples are collected and tested as follows: eacute phase - when symptoms are seen econvalescent phase - about 2 weeks after acute sample taken. Ua 4-fold increase in titer from acute to convalescent indicates a current infection.
32	Red Cell Suspensions Uprepare a 2% cell suspension using 0.5 mL washed packed cells. Final volume = (PCV) (100/desired %) Final volume = (0.5 mL) (100/2) Final volume = 25 mL so 25 mL - 0.5 mL = 24.5 mL of saline would be added to the 0.5 mL packed cells
33	QA versus QC Quality Assurance & Quality Control Definitions: eQuality assurance (QA): eQuality control (QC): the practice which encompasses all endeavors, procedures, formats, and activities directed toward ensuring that a specific quality or product is achieved and maintained. a component of QA which focuses on the analytic phase (procedures) of lab testing by monitoring overall reliability of results, including accuracy and precision, and comparing them to previously specified criteria.
34	Quality Assurance UQA programs may be designed according to the concept of Atotal quality management@ to include: equality control (QC) epolicies and procedures epersonnel training einspections and audits ecorrective action ecustomer satisfaction (Press-Gainey scores)
35	Quality Control UQC programs depend on specimens that are reproducible and are usually purchased from a supply company. UThese specimens are called Acontrols@ and the manufacturer supplies the acceptable range (" 2 SD) for each analyte. UTypes of controls include: ePositive and Negative controls eTri-level controls (High, Normal, Low) eNormal and abnormal controls UTypes of QC programs: eInternal or intralaboratory eExternal or interlaboratory
36	Accuracy vs Precision Accuracy: Precision: the closeness with which results agree with a known true value the closeness with which repeated results agree with each other Ayou can be precisely wrong@
37	The End of Safety & Quality Assurance Press the ESC key to end program