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LABORATORY TESTING: ITS ROLE IN DIAGNOSING AND MANAGING OCULAR

DISEASE

Tammy Pifer Than, MS, OD, FAAO

UAB School of Optometry

Birmingham, AL

tthan@uab.edu

Nothing to Disclose

Getting the Job Done...

External Lab TestingPCP

External laboratory

In-office samplingis it ok?

Before You Order Tests...

good case hx

narrow ddx

avoid “shot gun” approach

comprehensive ocular exam

If You Order Tests...

interpretLaboratory Tests and Diagnostic Procedures

6th edition – 12/2012Chernecky and Berger

– includes Herbal interactions

communicate treat refer

Getting the Job Done...

In Office Lab TestingPoint-of-Care

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CLIA Clinical Laboratory Improvement Act regulates all lab tests performed on

humans in US ensures quality laboratory testing “materials derived from the human

body… for the diagnosis, prevention or treatment…”

www.cms.hhs.gov/clia

CLIA

Classification FDA CertificatesCertificate of Waiver

~60% of 225,000 labs

Certificate of ComplianceCertificate of Accreditation

2 year renewals

CLIA can file for “Waived Status”Approximately 80 tests

random blood glucose

ESR

urine pregnancy test

RPS AdenoPlus

TearLab Osmolarity System

CLIA Certificate of Waiver

must meet criteria:enroll in CLIA programpay fee biennially ($150 for waived)follow manufacturers’ test instructions Good Laboratory Practices

“Educational” Visits2%/yearannounced

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Technology

• A rapid, point‐of‐care test that detects all known serotypes of Adenovirus

Test Components

Sterile Sample Collector

Test Cassette Buffer Vial

Survey – AAO 2013n = 340

Adenoviral Conjunctivitis

povidone-iodine ophthalmic solution (Betadine) (85:15)off-labeled use5% ophthalmic solution

Zirgan?

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Combined results from 7 eye care meetings held in 2013n = 649

In the works…

FST-100Foresight Biotherapeutics

0.1% dexamethasone and 0.4% povidone-iodine

Phase II trial

AL-46383A (N-chlorotaurine)Aganocide compound

Phase II trial

Doxovir

What’s Up and Coming?

www.clinicaltrials.govThe Food and Drug Administration

Amendments Act of 2007 (FDAAA or US Public Law 110-85) was passed on September 27, 2007

The law requires mandatory registration and results reporting for certain clinical trials of drugs, biologics, and devices

Dry Eye Disease and MMP‐9

Matrix metalloproteinases (MMP) are proteolyticenzymes that are produced by stressed epithelial cells on the ocular surface1

MMP‐9 in Tears

Non‐specific inflammatory marker

Normal range between 3‐41 ng/ml

More sensitive diagnostic marker than clinical signs1

Correlates with clinical exam findings1

Ocular surface disease (dry eye) demonstrates elevated levels of MMP‐9 in tears1

[1] Chotiakavanich S, de Paiva CS, Li de Quan, et al. Invest Ophthalmol Vis Sci 2009; 50(7): 3203‐3209.

Dry Eye Disease and MMP‐9

Increased concentrations of MMP‐9 can be found in other diseases or conditions, including:

Ocular rosacea

Meibomian gland disease

Sjögren’s syndrome

Corneal ulcers

Corneal erosions

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InflammaDry® Limit of Detection

Normal levels of MMP‐9 in human tears ranges from 3‐41 ng/ml

NEGATIVE TEST RESULTMMP‐9 < 40 ng/ml

POSITIVE TEST RESULTMMP‐9 ≥ 40 ng/ml

InflammaDry 4‐Step Process

* Release the lid after every 2‐3 dabs.  Allow the sampling fleece to rest along the conjunctiva for 5 seconds.

*

Reimbursement Strategy

CPT Code 83516 – Immunoassay for analyte other than infectious agent antibody or infectious agent antigen; qualitative or semiquantitative, multiple step method

$15.46

CLIA Waived Status

2/27/2014

Traditional Understanding of Sjögren’s

Sjögren’s is a chronic, systemic, progressive autoimmune inflammatory disease 

Characterized by the immune‐mediated (lymphocytic) destruction of the lacrimal and salivary glands

Early hallmark symptoms include dry eyes and dry mouth

Recent evidence suggests that all layers of tear film can be affected

Traditional Understanding of Sjögren’s

Primary Sjögren’s

Disease presents alone

• Secondary Sjögren’s

Subsequent to another autoimmune condition (e.g. rheumatoid arthritis)

It currently takes 4.7 years to receive an accurate diagnosis

Systemic effects are seen in 30‐70% of patients

Myths of Sjögren’s

“All Sjögren’s patients are identified and diagnosed”

“There are only a few patients in my practice”  

“Nothing can be done for the patients if they are diagnosed”

“Sjögren’s Syndrome does not have serious long‐term consequences, it is just a nuisance”

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Sjögren’s Syndrome and Non‐Hodgkin’s Lymphoma

Solans‐Laque R, et al, Seminars Arthritis Rheum, 2011

Lymphoma risk

~10% at 15 years

Risk  with time 

Ioinnidis et al, Arthritis Rheum, 2002

Primary SS

20% deaths due to lymphoma

Mortality in pSS

Ioinnidis et al, Arthritis Rheum, 2002

723 patients studied in Greece

1 in 5 deaths of pSS patients due to lymphoma

Early detection is key!

Markers for Sjögren’s

• The classical serological markers for Sjögren’s are anti‐Ro/SS‐A and anti‐La/SS‐B antibodies• Other antinuclear antibodies (ANA) and rheumatoid factors (RF) are also included as the more common serological markers detected

• The combined serology sensitivity and specificity of the classical markers is around 40‐60%• None of the currently recommended serology tests diagnose Sjögren’s early in the disease progression

• In approximately 20‐30 % of cases no classic Sjögren’santibodies are found

New Markers

Salivary Gland Protein 1 (SP‐1) Submandibular and lacrimal glands

Carbonic Anhydrase 6 (CA6) Involved in buffering capacity of saliva

Submandibular and parotid glands

Parotid Secretory Protein (PSP) Involved with binding and helping to clear various infections

The Specifics

No CLIA waiver

Saturate at least 3 of 5

CPT 36416Collection of blood by capillary blood

specimen (e.g. finger, heel, ear stick)

Insurance Info

Call FedEx

IMMCO Lab Only

OR…

Microbiology

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Cultures and Sensitivities

specimen preparation is important no anesthetic – if possible sterile swab plate onto culture media culturette media:Thioglycolate brothBlood agarChocolate agarSaboraud’s agar

Transport Media

Amies media without charcoalHigher yield than other media

Comparable to plates

Subconjunctival Hemorrhage

Historyfrequency

medications

activity

Examination

Subconjunctival Hemorrhage

Idiopathic Valsalva maneuver HTN, DM Von Willebrand’s Disease 1-2%

Severe hepatic disease Leukemia Vitamin K deficiency AIDs

Subconjunctival Hemorrhage

Blood pressure CBC with differential PT (prothrombin time) PTT (partial thromboplastin time) INR (international normalized ratio)

Prothrombin Time (PT)

prothrombin:vitamin-K dependent glycoprotein produced

by liverneeded for firm fibrin clot formation

PT – measures time for clot formationreagent tissue thromboplastin and calcium

are added to citrate plasma

avoid coffee and alcohol for 24 hours before test ↓ time

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Prothrombin Time (PT)

each lab has normal value normal range is 2 secs Adult 12-14 sec International Normalized Ratio (INR)standardizes PT resultsINR = (Patient’s PT in seconds)ISI

Mean normal PT in secondsISI = international sensitivity indexNormal 0.9 – 1.3Coumadin therapy

Partial Thromboplastin Time (PTT) evaluates how well coagulation

sequence is functioning time for recalcified, citrate plasma takes

to clot after partial thromboplastin is added

Activated PTTcommercial activating materials used to

standardize the testcurrent method of the test

Standardized times reported by each lab< 35 seconds

Coagulation Studies

recurrent subconjunctivalhemorrhages

non-traumatic hyphema

± artery or vein occlusion

pre-op ocular surgery

To Treat or not to Treat.

34 YOWF

CC: HAs, double vision, dizzy

OHx: no trauma, LEE – long time ago

MHx: Voltaren, Zantac

Magnetic Resonance Venography(MRV)

Emerging imaging toolVeins of abdomen, pelvis, thorax and

extremities

Duplex sonography hindered by acoustic access

DVTNew gold standard

Cerebral Venous Sinus Thrombosis

Causes increased intracranial pressure

Can be life-threatening

Should be a DDX for every case of IIH

CVST9-26% of patients

Reference Lin A, Foroozan R, et al, Occurrence of cerebral venous

sinus thrombosis in patients with presumed idiopathic intracranial hypertension. Ophthalmology 1996; 113(12);2281-84.

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Management

weight loss

acetazolamideDiamox Sequels

steroids??

ON sheath decompression

LP shunt

Before you prescribe Diamox

baseline electrolytes

CBC with differentialR/O blood dyscrasias

monitor every 6 months

Electrolytes Na+

135.0 – 145.0 mmol/L

K+

3.50 – 5.3 mmol/LPanic Level: <2.5 or >6.6

Cl-

97 – 107 mmol/LPanic Level: <80 or >115

CO2 total content blood

21.0 – 31.0 mmol/L Increasedalcoholismairway obstructionpneumoniadrugs (e.g. antacids)

Decreaseddehydrationacetazolamide

measures compliance - < 20 mEq/L

tetracyclines

SMA-6

Sequential multiple analyzer (SMA) automated system that analyzes multiple

blood values from one tube of blood SMA-6Carbon dioxideChlorideCreatininePotassiumSodiumUrea nitrogen

SMA-7

Carbon dioxide

Chloride

Creatinine

Glucose

Potassium

Sodium

Urea nitrogen

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SMA-12AlbuminAlkaline phosphataseAspartate aminotransferaseBilirubinCalciumCholesterolGlucoseLactate dehydrogenasePhosphorusProteinUrea nitrogenUric acid

Glad you looked!

58 year old female

CC: SpRx broken

OHx: unremarkable

MHx: unremarkable, no meds

20/20 OD; 20/20 OS

Random Blood Glucose

note when patient ate laste.g. 220 mg/dL pp 3 hours

pp = post-prandial

diabetic if: 200 mg/dL with symptoms

can do in-office

encourage patients to do this!

± reimbursed (CPT code – 82962)

Fasting Plasma Glucose

no food or drink for 8-12 hours

diabetes if 126 mg/dLmust repeat if asymptomatic

IFG = 100 – 125 mg/dL

also increased with:steroids

stress

diuretics

Oral Glucose Tolerance Test (OGTT)

75 g oral glucose

check urine and blood at intervals

non-diabetic BS will return to fasting levels in 3 hours

diabetic if 200 mg/dL at 2 hours

impaired GT if 140 and < 200 mg/dL at 2 hours

not needed if FBS > 200 mg/dL

Glycosylated Hemoglobin HbA1c

checks long-term control glycosylated HgB stays with RBC for its

entire life normal = 4.3-6.1%Diabetic if 6.5%

diabetic goal < 7.0% ask patients! A1C Now InView multitest system CPT 83036QW

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A1C Now InView CLIA waived

$129.00 for 10 tests

Medicare Coverage$13.56

Private insurance$18.00

Glycoslyated HemoglobinA1C Blood Glucose Levels

12% 345 mg/dL

11 310

10 275

9 240

8 205

7 170

6 135

5 100

4 65

1% A1C = 30 mg/dL

eAG

estimated Average GlucoseA1C-derived average glucose study

Diabetes Care 2008 31(8);1704-7.

Linear relationship

Recommended by ADA

eAG = 28.7 * A1C – 46.7

Pre-Diabetes

IFG or IGT OR A1C 5.7-6.4% Weight loss of 7% of body weight 150 minutes/week of moderate activity

www.diabetes.org

Executive Summary – Standards of Medical Care in Diabetes - 2014

ESR

erythrocyte sedimentation rate

nonspecific test for inflammation

mm/hr

M: age/2

F: (age+10)/2

usually > 60 mm/hr in GCANormal in 7-20%!!!

C-Reactive Protein (CRP) abnormal serum glycoprotein produced by

liver during acute inflammation disappears rapidly once inflammation

subsides 4 hour fast from food/fluids alternative to ESR more informativeESR high in most elderlyno cross interference

normal: Qualitative – negative; Quantitative – 0-10 mg/L

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The Observant Patient..

52 YOWM

CC: “inferior vision OS is dim”

MHx: diabetic x 20 years; poorcontrol

VAs: OD 20/20 OS 20/20-2

LEE: 6 month priortwo dot hemorrhages OD

?????

Causes of Optic Nerve Edema

Arteritic Ischemic Optic Neuropathy Nonarteritic Ischemic Optic Neuropathy Central Retinal Vein Occlusion Compressive Optic Nerve Head Tumor Diabetic Papillopathy Infiltration of Optic Nerve Head Malignant Hypertension Papilledema Papillitis Papillophlebitis Thyroid Ophthalmopathy

Thyroid Testing

Sensitive Thyroid Stimulating Hormone0.3 – 3 μU/L

↓ - hyperthyroidism

↑ - hypothyroidism

Also if:Proptosis

SLK

Etc.

Lipid Panel

12 hour fast

total cholesterol

LDL

HDL

triglycerides

risk for CAD

ratio total cholesterol / HDL

Cholesterol

over half of adults in US have cholesterol > 200 mg/dL

desirable: 160-200 mg/dL borderline: 200-239 mg/dL high 240 mg/dL Outside US cholesterol x 0.0259 mmoles/L (international

units)200 mg/dL = 5.18 mmol/L

More Numbers…

HDLgood 35 mg/dL

women probably 45 mg/dL

1 mg/mL risk of HD 2-3%Helsinki Heart Study (gemfibrizol in men )

LDLgood < 130 mg/dLhigh 190 mg/dL

Ratio (Total / HDL) 5:1

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Triglycerides

normal < 200 mg/dLwomen probably < 150

borderline 200-400

high 400-1000

very high > 1000

Vertical Auto Profile (VAP) Test

More detailed info

Beginning to replace lipid profile

Only test to meet ADA and American College of Cholesterol Guidelines

Subclasses of lipoproteins

Some additional lipids measured

VAP

Total VLDL

HDL + LDL + VLDL (sum total cholesterol)

LDL + VLDL (total non-HDL)

Total apoB100 (apolipoprotein B100)

Lp(a) cholesterolInherited risk factor for atherosclerosis

IDL – intermediate density lipoprotein↑ in FHx of diabetes

VAP

LDL-RCLDL bound to C-reactive protein

Found at site of atherosclerotic plaques

Lp(a)+IDL+LDL (sum total LDL-C)

LDL size patternA

A/B

B – 4X greater risk of heart disease than A

VAP

HDL-2Particularly protective

HDL-3

VLDL-3Proposed that if ↑, more likely to develop

diabetes

Additional Testing…

ImagingMRI – imaging of choice

Gadolinium– Nephrogenic Systemic Fibrosis

CT – caution if ordering contrast

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CT Contrast

Iodineenhances visibility of vascular lesions

Administered IV (or intrathecal)

1:40,000 incident of AE

BUN and Creatinine

NPO

Good medication hxd/c Metformin (Glucophage) prior to

procedure

CI if shellfish allergy

BUN (Blood Urea Nitrogen)

actually performed on serum or plasma12% higher than blood

nitrogen portion of urea urea is formed in liver from protein

breakdown filtered through renal glomerulismall amount reabsorbed in the tubulesremainder excreted in urine

azotemia – elevated BUNnonspecific prerenal, renal, or postrenal

BUN (Blood Urea Nitrogen)

must be compared over time or evaluated with other testsrenal function – also assess creatinine levels

fasting not required Adult 5-20 mg/dL >60 8-21 mg/dL increased BUNmany conditions and many drugs

decreased BUNalcohol abuse, diet lacking protein, liver

destruction, late pregnancy

CREATININE

product of anaerobic energy-producing creatine-phosphate metabolism in skeletal muscle

excreted by kidneysincreased levels indicative of decreased

glomerular filtration rate

Avoid excessive exercise for 8 hours and avoid excessive red meat for 24 hours before testing

CREATININE

Normalfemales 0.5 – 1.1 mg/dLmales 0.6 –1.2 mg/dLelderly – may be lower

Creatinine clearance, urine24 hour collection

Creatinine clearance, serum - urine6, 12, or 24 hour collectionblood sample collected anytime during urine

collection period Creatinine with eGFR

Diabetic Papillopathy

0.4 – 2% of diabetics

characteristicssectoral or total ON edema

± peripapillary hemorrhages

± nerve fiber layer infarcts

± macular edema

unilateral or bilateralasymmetric

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Diabetic Papillopathy

retinopathy does not need to be present

small optic nerve cupping 0.3/0.3

prognosis:significant or complete recovery in several

months

may have residual pallor and VF defect

pathophysiology is unclear

Laboratory Testing for Uveitis

ACE

angiotensin converting enzyme

produced by a variety of cells including granulomatous cells

best for patients > 20 YO

helps confirm dx of sarcoidosisACE elevated in 60%

12 hour fast

Sarcoidosis

Laboratory TestingChest X-Ray

Serum Angiotensin Converting Enzyme (ACE)

Conjunctival or lacrimal gland biopsy

Serum lysozyme

Serum calcium

Gallium scanNuclear medicine test

Radioactive gallium citrate is injected

Hot spots at site of inflammation

ANA

antinuclear antibody

evaluates immune system

8 hour fast

screening test for SLE

sensitive but not specific

Non-specific testRheumatoid arthritis

Scleroderma (60-90%)

Sjögren syndrome

ANA

Look at staining patternHomogenous (SLE)

Speckled (SLE, Sjögren syndrome, scleroderma, etc.)

Peripheral or rim (SLE)

Nucleolar (Sjögren syndrome, scleroderma)

normal: nonreactiveTiter <1:20

results in 4-5 days

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ANA - Sensitivity

SLE- 95 %

Scleroderma- 60-90%

Rheumatoid arthritis- 41%

Sjőgren syndrome- 48%

JIA with uveitis – 80%

HLA-B27

Human Leukocyte AntigenSurface of WBC

Immune function

HLA B-27 present in up to 8% of population50-60% with acute anterior uveitis

Normal: negative

HLA-B27

Order if acute, recurrent, unilateral anterior uveitis50-80% are seronegative spondyloarthropathies

Ankylosing spondylitis

Reactive arthritis (FKA Reiter’s syndrome)

Inflammatory bowel disease

Psoriatic arthritis

p-ANCA perinuclear-Antineutrophil Cytoplasmic

Antibody

Autoimmune antibodies directed against the lysosomal enzymes in neutrophil granules

p-ANCAAntibodies found near the nucleus

(perinuclear)SLE, Rheumatoid arthritis, ulcerative colitis

Elevated in Crohn’s disease

C-ANCA c-ANCA (classical)Antibodies found scattered in the cytoplasm of

neutrophils

Only test available for the diagnosis of vasculitis

Granulomatosis with polyangiitis(Wegener’s)

Normal: negative; titer <1:40

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Rheumatoid Factor

Positive titers in numerous collagen vascular diseases

(+) in 70-80% of patients with Rheumatoid Arthritis

Also (+) with:SLE, Sjogren, TB, sarcoid, viral infection

Negative finding most useful (<1:20)JIA

Rheumatoid Arthritis

Rheumatoid Factor

Anti-cyclic citrullinated peptide antibodyanti-CCP

Auto-antibody frequently seen in RA

Allows early dx

Syphilis: Ever Had It?

FTA-ABSfluorescent treponemal antibody absorption

test

ordered more frequently

positive even after treatment

MHA-TPmicrohemagglutination treponemal pallidum

test

Syphilis: Do you have it now?

RPRrapid plasma reagin test

VDRLvenereal disease research laboratory test

EIAEnzyme immunoassay

Other Testing in AU

PPDCheck in 48-72 hours

Positive for active and inactive TB

Lyme titers

Etc.