MGH Pathology Service Laboratory Reflex Test Protocols 2016

Reflex Test Protocols  Reflex tests fall into 2 categories: standard industry practice (example: sensitivities, interpretations, and confirmations) and institutional practice based on staff clinical practice  Tests that are reflexed should be reasonable and medically necessary. The medical community should approve reflex protocols.  The physician must have the option of selecting a reflex protocol or a single test.

Blood Transfusion Services Test ordered on Requisition Initial Test Performed Criteria for Reflex Test Ordered by Reflex Screen Antibody Screen Positive screen Antibody identification Cold agglutinin screen Cold agglutinin screen Positive (>1+) Cold agglutinin titer Direct antiglobulin test Direct antiglobulin test IgG positive and transfused w/in 3 Eluate mo Fetal Screen Fetal screen Positive Kleihauer-Betke

Core Lab - Chemistry Test ordered on Requisition Initial Test Performed Criteria for Reflex Test Ordered by Reflex Thyroid Screen TSH TSH>5.00 Free T4 TSH<0.40 Free T4. T3 Monitoring L-T4 Therapy TSH TSH<0.05 Free T4 Thyroglobulin Thyroglobulin Specimen received and test Thyroglobulin requested Direct LDL Lipid Panel with calculated LDL >399 Direct (measured LDL) Free testosterone Total testosterone, sex hormone Albumin <3.5 Free testosterone measured by binding globulin (SHBG), albumin, equilibrium dialysis free testosterone (calculated), % free testosterone (calculated).

Notes: The albumin is needed for the calculation but will not be reported. All other values will be reported. If the total testosterone is outside of the measurement range, free testosterone and % free testosterone will not be reported.

Core Lab - Emergency Department Test ordered on Requisition Initial Test Performed Criteria for Reflex Test Ordered by Reflex Urinalysis Macroscopic urinalysis 1+, 2+, 3+ for blood protein, Microscopic leukocytes Troponin-l Troponin-l Borderline or positive Troponin-T Rapid Influenza screen Rapid flu screen Admission samples Confirmation in main Microbiology Rapid Influenza screen Rapid flu screen Negative on Pediatric case and at Viral Panel in main Microbiology request of MD Rapid Strep A Antigen Strep screen Presumptive negative Confirmation on second swab in main Microbiology

Cytology Test ordered on Requisition Initial Test Performed  Criteria for Reflex Test Ordered by Reflex Pap Smear Liquid-based PAP Atypical cytology HPV test Anal Pap Smear Liquid-based PAP Atypical cytology HPV test

Diabetes Test ordered on Requisition Initial Test Performed Criteria for Reflex Test Ordered by Reflex Hemoglobin A1C Hemoglobin A1C >4.50% or <14.50% Calculated mean blood glucose

MGH Laboratory Reflex Test Protocols 2016 – Approved by Medical Policy 2016 MGH Pathology Service Laboratory Reflex Test Protocols 2016

Health Centers – Chelsea and Revere Test ordered on Requisition Initial Test Performed Criteria for Reflex Test Ordered by Reflex Urinalysis UA-Macroscopic Any of the following Urine Sediment Protein:1+ or more Blood: Trace or more Leukocyte esterase: Positive CBC MCHC>38.0 Manual hematocrit Differential – Manual Differential – Manual - Clinical requests pathologist review - Blasts seen - Unusual abnormal white cells Rapid Influenza screen Rapid flu screen Admission samples Confirmation in main Microbiology Rapid Influenza screen Rapid flu screen Negative on Pediatric case and at Viral Panel in main Microbiology request of MD Rapid Strep A Antigen Strep screen Presumptive negative Confirmation on second swab in main Microbiology

Core Lab - Hematology Test ordered on Requisition Initial Test Performed Criteria for Reflex Test Ordered by Reflex Hemoglobin electrophoresis Hemoglobin fractionation Test performed Pathologist review of results. Pathologist review of peripheral blood smear if abnormal findings seen on hemoglobin electrophoresis. No concurrent CBC CBC Sickle screen Sickle screen Previous positive Test result Hemoglobin electrophoresis Urinalysis Macroscopic urinalysis Positive WBC screen occult blood Microscopic or >TRACE albumin (i.e., 1+ or more) CBC CBC Instrument flag PLT review on smear Manual PLT first time <50,000 Platelet Instrument flag Manual hematocrit; other RBC RBC > 7.0 million/uL parameters not reported MCHC > 38.0 Manual hematocrit Differential Differential-automated Instrument flag Differential or automated differential with manual review Differential-manual Differential-manual  Clinician request Pathologist review  Blasts seen for first time on the patient  Unusual abnormal white cells PT PT  >138 and/or reason to Manual PT (and fibrinogen when query accuracy of result e.g. initial PT >138) low fibrinogen  interfering substance (lipemia, gross hemolysis)  Instrument flag

MGH Laboratory Reflex Test Protocols 2016 – Approved by Medical Policy 2016 MGH Pathology Service Laboratory Reflex Test Protocols 2016

Hematology Pathology Test ordered on Requisition Initial Test Performed Criteria for Reflex Test Ordered by Reflex Bone Marrow Exam Specimen received and test Pathologist review of blood smear request (from CBC ordered <24 hours prior to bone marrow sample or slide from CBC with manual differential/special slide ordered < 7 days prior to bone marrow) Flow cytometry for hematologic Specimen received and test CBC with differential, special slide malignancy request and pathologist review. CSF Flow Cytometry Ordered  CSF Total Nucleated  TNC> 5/µL or history of CSF flow cytometry Cell Count (TNC) hematologic malignancy  Chart review for history  Clinician requests policy (If reflex criteria not met, flow of hematologic override cytometry will not be performed by malignancy default, but clinician may request override) Lymphoma staging bone marrow  Morphologic  Pancytopenia or Routine cytogenetics examination by a question of MDS hematopathologist  Flow, morphology and Cytogenetics with stimulation  Flow cytometry or IHC positive for  IHC as needed lymphoma  No evidence of marrow Cytogenetics not performed (test involvement by cancelled by Hematopathology). lymphoma based on flow, morphology or IHC Bone marrow examination for  Morphologic Diagnosis Molecular Add-on tests suspected acute leukemia examination by a hematopathologist AML SNaPshot, FLT3, NPM1, CEBPA  Flow cytometry  IHC as needed APML SNaPshot, FLT3, PML-RARA

B-ALL BCR-ABL qualitative

MDS SNaPshot

MPN or MDS/MPN overlap JAK2 V617F and SNaPshot

CMML SNaPshot, NPM1, FLT3

Other Discuss with clinician

MGH Laboratory Reflex Test Protocols 2016 – Approved by Medical Policy 2016 MGH Pathology Service Laboratory Reflex Test Protocols 2016

Immunology Test ordered on Requisition Initial Test Performed Criteria for Reflex Test Ordered by Reflex Protein electrophoresis on serum Protein electrophoresis on Presence of 1 or more abnormal Immunofixation to characterize or CSF agarose gel bands in the gamma globulin zone abnormal bands with physician review Protein electrophoresis on serum Protein electrophoresis on Presence of abnormal banding Immunofixation to characterize or CSF agarose gel and altered immunoglobulin profile abnormal bands and, as needed, free kappa and lambda light chains by nephelometry with physician review Protein electrophoresis on serum, Protein electrophoresis on Test performed Physician review urine, CSF agarose gel Urine Bence Jones proteins Agarose gel electrophoresis and Abnormal band on agarose gel Immunofixation to characterize immunoelectrophoresis not identified by abnormal bands and, as needed, immunoelectrophoresis free kappa and lambda light chains by nephelometry with physician review Antinuclear antibody ANA screen Presence of staining of substrate ANA Titre and pattern Physician nuclei or cytoplasm review Quantitation of serum IgG, IgA, IgM by nephelometry If lgG>7 g/dL, lgA >3 g/dL, or lgM Viscosity immunoglobulins >5 g/dL Quantitation of serum lgG lgG by nephelometry <500 mg/dL and adult patient Immunofixation electrophoresis, and, as needed, free kappa and lambda light chains by nephelometry and physician review of findings Cryocrit Cryoprecipitable protein Cryoprecipitable protein present If not previously tested within 6 months to 1 year, cryoprecipitable protein is identified by immunodiffusion, immunofixation, and physician review ENA panel ELISA If ELISA positive for any antigens Perform ANA test on Hep2 cells if (Ro/La/Sm/RNP/Jo/Sci-70) in ENA panel not already performed within last 2 months

MGH Laboratory Reflex Test Protocols 2016 – Approved by Medical Policy 2016 MGH Pathology Service Laboratory Reflex Test Protocols 2016

Microbiology Test ordered on Requisition Initial Test Performed Criteria for Reflex Test Ordered by Reflex Routine culture on CSF Culture Cloudy fluid Gram stain Lower respiratory culture Culture Evaluation of specimen quality Gram stain Lower female tract culture Culture Diagnosis of bacterial vaginosis Gram stain made by gram stain not culture. Culture performed for yeast and GC Gram stain Gram stain Presence of potential pathogens TB, fungal or anaerobic cultures observed and/or smears Brain abscess Routine culture Specimen site Anaerobic culture Gram stain routine culture Bacterial cultures Routine Culture  Isolation of clinically significant  Susceptibility testing organisms requiring  Vancomycin MIC to check for susceptibility VISA or VRSA  Isolation of methicillin resistant  Confirmatory test for the Staph aureus from blood, body presence or absence of the fluid, or wound MecA gene or the gene product  Staphylococci from blood, body PBP 2a for certain fluid, or wound that require staphylococci confirmation of methicillin test  Confirmatory test for presence  Isolation of certain gram of extended-spectrum B- negative bacilli resistant to 3rd lactamases generation cephalosporins Mycobacterial culture Mycobacterial culture Isolation of clinically significant Susceptibility testing organisms Mycobacterial culture from Mycobacterial culture Body site AFB smear normally sterile body site AFB Mycobacterial culture Test order AFB smear HSV Culture on CSF HSV PCR Direct fluorescence antibody DFA for both HSV and VZV DFA order for either HSV or VZV DFA for both HSV and VZV detection (DFA) for Herpes simplex virus (HSV) or Varicella zoster virus (VZV) Lyme serology Lyme serology by EIA Positive Immunoblot IgG and IgM Syphilis antibody screen (ELISA) Syphilis antibody screen (ELISA) Positive Syphilis antibody screen RPR, RPR titer, alternative (ELISA) treponemal specific testing if needed Ova & Parasites or Blood O&P or Blood Parasite Isolation of clinically significant Parasite identification parasites parasite Parasite count Hatching test Viral antigen Respiratory antigen testing for 5 Positive for specific virus Any one of the following: viral antigens via indirect  Adenovirus fluorescent antibody staining  Parainfluenza 1,2,3  Human metapneumovirus Viral respiratory culture (Non- Respiratory antigen testing for 5 Antigens are recommended tests All of the following listed above. tissue specimens) viral antigens via indirect fluorescent antibody staining AND

Influenza and RSV PCR PCR are the recommended tests Influenza and RSV

Complete viral culture on non- Viral culture and CMV shell vial Shell vial culture is recommended CMV shell vial culture respiratory tissue culture test HSV(PCR) Pathologist review with M. pneumoniae (PCR) interpretation Hep C Virus (PCR) Hepatitis C genotyping HIV serology and viral load Pneumocystis exam Viral hepatitis serology Lyme immunoblot Anaerobic culture Anaerobic culture Gram stain Gram stain results dictate Gram stain additional media for optimal organism recovery Routine culture on specimen from Culture Specimen obtained surgically and Gram stain OR coming from the OR

MGH Laboratory Reflex Test Protocols 2016 – Approved by Medical Policy 2016 MGH Pathology Service Laboratory Reflex Test Protocols 2016

Culture of blood or normally sterile Routine culture for yeast and Yeast growth Antifungal susceptibility on yeast sites bacteria isolates Blood culture Blood culture followed by gram Presence of gram-positive or Multiplex molecular test for stain if growth is detected negative bacteria or yeast organism identification and detection of resistance markers Stool examination for O&P (Ova Direct immunofluorescent assay Submission of completed patient Permanent stained smear, and Parasites) for Giardia and Cryptosporidium history form concentration with wet mount or NAAT for Giardia, Cryptosporidium and Entamoeba histolytica HCV Genotyping (added 1/06) No Hepatitis C Viral Load HCV Quantitative Viral Load performed at MGH within previous 30 days. HIV Genotyping (added 1/06) No HIV viral load performed at HIV Quantitative Viral Load MGH within previous 30 days Malaria Screen BINAX rapid screen Positive BINAX test Thick and thin smear for confirmation, parasite burden, and speciation Clostridium difficile toxin assay Combination assay for GDH and Indeterminate result PCR assay for toxigenic C. difficile Toxins A&B •Ehrlichia-PCR (or synonym) Ehrlichia/ Anaplasma molecular NA NA •Anaplasma-PCR (or synonym) detection •Ehrlichia/ Anaplasma PCR (or synonym) Herpes simplex virus (HSV) HSV DFA Negative HSV DFA HSV culture culture or direct fluorescence antibody detection (DFA) Influenza PCR or Respiratory PCR for Influenza A, Influenza B Test order for any of these viruses PCR for Influenza A, Influenza B syncytial virus (RSV) PCR and RSV and RSV will performed in all cases Nucleic acid amplification test NAAT for both Chlamydia NAAT test order for either NAAT for both Chlamydia (NAAT) for Chlamydia trachomatis trachomatis and Neisseria Chlamydia trachomatis or trachomatis and Neisseria or Neisseria gonorrhoeae gonorrhoeae Neisseria gonorrhoeae gonorrhoeae

Molecular Pathology Test ordered Initial Test Performed Criteria for Reflex Test Ordered by Reflex A pathologist review and written interpretation accompany all results of molecular pathology tests.

MGH Laboratory Reflex Test Protocols 2016 – Approved by Medical Policy 2016 MGH Pathology Service Laboratory Reflex Test Protocols 2016

Special Test ordered on Requisition Initial Test Performed Criteria for Reflex Test Ordered by Reflex Screen Positive Screen confirmation; anticardiolipin antibody added even if lupus anticoagulant is negative. Factor VIII, IX, XI to rule-out factor inhibitor causing false positive results Protein C Protein C functional <70% activity Antigenic Protein C, Factor VII if indicated to distinguish between hereditary and acquired etiologies. Antithrombin III Functional Antithrombin III <70% activity Antigenic Antithrombin III Protein S Functional Protein S <70% activity Free Protein S antigen, fibrinogen and functional FVIII activity Activated protein C resistance or Activated protein C resistance ≤2.1 Leiden by DNA assay Factor V Leiden Lupus anticoagulant Prolonged PT Evaluation (mixing PT, removal of if PTT also Mixing study normal, prolonged, Factor assays, lupus studies) prolonged, mixing study or "fades" anticoagulant, and/or factor inhibitor tests if indicated Prolonged PTT Evaluation (mixing PTT, removal of heparin, mixing Mixing study normal, prolonged, Factor assays if mix is normal; studies) study or "fades" lupus anticoagulant if mix is prolonged, factor VIII if mix "fades"; all three tests if mix results inconclusive; factor inhibitor tests if indicated Multiple individual As ordered Patient not on coumadin or other If missing a test from the usual hypercoagulation tests reason for not performing tests. screen (activated protein C resistance, protein C, protein S, antithrombin, Lupus anticoagulant, Anticardiolipin, prothrombin G20120A), it will be included Reptilase time Reptilase time > 24 seconds Fibrinogen Degradation Products (or D Dimer), Fibrinogen Factor V, factor II, or As ordered The medical record indicates the The appropriate test will be clinician wanted factor V Leiden, performed as indicated by the prothrombin G20210A ("factor II"), patient's medical record or a chromogenic factor X or anti- factor Xa A written pathologist's interpretation accompanies all results of complex coagulation tests, including mixing studies, hypercoagulation studies, antiphospholipid antibody studies, factor assays, von Willebrand tests, aggregation studies, and in some cases anti-factor Xa assays (heparin, low-molecular weight heparin, and Fondaparinux drug levels).

MGH Laboratory Reflex Test Protocols 2016 – Approved by Medical Policy 2016