1. Clinical Utility of Rheumatologic Tests: A Guide to Interpretation
Sheetal Desai MD MSEd

2. Questions for you…
1. What is the most likely diagnosis of a patient with a positive ANA?
2. What is the most common cause of an elevated ESR >100?
3. A preferred screening lab test has a high sensitivity or high specificity?
4. Which of the following lab tests is most specific? ESR or CRP

3. Common Misperceptions
Positive result = Disease
These tests are free, so order as many as you want
“money grows on
trees”

4. Rheumatology Lab Tests
Anti-CCP
ANCA
CRP s
anti-Smith
ANA
dsDNA s RF

5. Rheumatology Lab Tests
Are relatively young
Have varied sensitivity and specificity
Little value as a screening test
Blind ordering can lead to diagnostic confusion

6. Clinical Scenario: Case 1
68 year old patient presents to the ER with fevers, chills and general malaise for one week.
PMH: DM, HTN
Meds: Metformin, Januvia, Norvasc, Lisinopril
NKDA
T 38.6 in ER, BP 110/60, HR 108
PE: normal
Labs: ESR 90, CRP 10mg/dl, WBC 10, Hg 12, Plt 450,000, Cr 1.1, AST 30, ALT 28
Ddx?

7. Acute Phase Proteins
Proteins that show an increase >25% with inflammation
Synthesized in the liver
Group 1- increases by 50%: C3 and ceruloplasmin
Group 2- increases 2-4 fold: fibrinogen, haptoglobin, alpha 1 antitrypsin, alpha 1 chemotrypsin, alpha 1 glycoprotein
Group 3- increases by several hundred fold: CRP, serum amyloid A

8. Erythrocyte Sedimentation Rate (ESR)
What exactly is it???
Indirect measure of acute phase protein fibrinogen
Measured by the Westergren method
Vertical column mm long
ESR = the distance the RBC travel in one hour

9. What is a normal ESR??? At UCI 0-20 is normal
However ESR increases with age and is increased in women
For Men: age/2
For Women: (age + 10)/2

10. Factors that Increase ESR
Inflammatory disease
Infections
Malignancy
Increase in globulin proteins
End Stage Renal Disease
Extensive Tissue Necrosis
Pregnancy
Age

11. Factors that Decrease ESR
Elevated plasma viscosity
Increased RBC’s- Polycythemia
Abnormal RBC shape- Sickle Cell
Hepatic Necrosis
Hypofibrinoginemia
Congestive Heart Failure
Extreme Leukocytosis
Trichinosis

12. Etiology of ESR >100
#1 Infection
#2 Malignancy
#3 Rheumatologic

13. How to Use ESR Not very sensitive or specific
Use as diagnostic criterion only for Temporal Arteritis (TA) and PMR
Useful in monitoring PMR, TA, RA
Can be useful for monitoring disease course and treatment response
Extreme elevations in the ESR rarely occur without evidence of serious disease

14. C Reactive Protein (CRP)
What does the C stand for?
C-polysaccharide from the pneumococcus cell walls
Acute phase protein- Group 3
Exclusively produced by hepatocytes
Direct measure of inflammation

15. C Reactive Protein (CRP)
Normal Levels <1
Moderate elevation
See in most of the rheumatic conditions- RA, SLE, Sjogren’s
Marked elevation >10
See in serious bacterial infections, severe RA, Vasculitis, PMR

16. C Reactive Protein (CRP)
Levels rise within hours of stimulus
Peaks within 2-3 days
Half life 8 hours
With effective treatment of the underlying cause, levels can normalize within hours

17. CRP vs. ESR Rises quickly Falls quickly Direct marker of inflammation
Narrow range of results
High sensitivity
High specificity
High reproducibility
NOT affected by factors (age, gender, anemia, RBC shape, plasma proteins)
Rises slowly
Falls slowly
Indirect marker of inflammation
Wide range of results
Mod sensitivity
Mod specificity
Mod reproducibility
Affected by many factors

18. Clinical Scenario: Case 1
68 year old patient presents to the ER with fevers, chills and general malaise for one week.
PMH: DM, HTN
Meds: Metformin, Januvia, Norvasc, Lisinopril
NKDA
T 38.6 in ER, BP 110/60, HR 108
PE: normal
Labs: ESR 90, CRP 10mg/dl, WBC 10, Hg 12, Plt 450,000, Cr 1.1, AST 30, ALT 28
Ddx? Ur Cx and Blood cx Gram neg rods

19. Clinical Scenario: Case 2
42 year old caucasian female, otherwise healthy, comes to clinic complaining of fatigue. She complains of fatigue, poor sleep habits and aches and pains over the past year. Her joints have been bothering her, especially her hands. There is a discomfort and stiffness that comes and goes, and usually involves one hand at a time. She states that at times her hands have been mildly swollen and have limited her function.

20. Clinical Scenario Vitals T 99.2, BP 116/80, P 90 R 16 98%RA
PE is unremarkable
HEENT: WNL, no rash
CV: RRR, no murmurs
Pulm: CTA bilaterally
Abd: benign, no organomegaly
Ext: no edema, FROM of all joints, no appreciable joint swelling in wrist, MCP, PIP, DIP joints. No deformities. No rash

21. Clinical Scenario Lab Tests: CBC: WBC 6.3, Hg 12.7, Plt 266
Electrolytes: Na 138, K 4.2, Cr 0.7
TSH 3.8 (normal )
RF: negative
ANA: positive, titer 1:80

22. ANA What exactly are they?
Antibodies that bind to various antigens in the nucleus of a cell
How is it measured?
Indirect Immunofluorescence

23. Antinuclear antibodies
Indirect Immunofluorescence Assay
Take patient serum and add it cells
If there are antibodies they will bind
Add a fluorochrome tag
View under a fluorescent microscope
If it lights up in then positive 1:40
Dilute sample and repeat, 1:80, 1:160, 1:320, 1:640, 1:1280, etc

24. Antinuclear Antibodies

25. Antinuclear Antibodies
Staining Patterns
Observer dependent
Not sensitive
Not specific
Only LOOSELY associated with certain disease states

26. Antinuclear Antibodies
What does the staining pattern mean?
Homogenous SLE
Rim SLE
Speckled Sjogren’s, MCTD
Diffuse nonspecific
Nucleolar Scleroderma
Anti-centromere CREST

27. Antinuclear Antibodies

28. Positive ANA
What disease states do you see it?

29. ANA associated Diseases
Rheumatic Conditions
Auto- Immune
Misc
Lupus
Polymyositis
Grave’s
Aging
Drug-induced Lupus
Dermato- myositis
Primary Biliary Cirrhosis
Primary Pulmonary Hypertension
Scleroderma RA
Hashimoto Thyroiditis
Sjogren’s
Vasculitis
Autoimmune Hepatitis
MCTD

30. ANA with age
For every year after age 50, percentage of ANA positivity increases 1%/year
For example
Age %
Age %
Age %

31. Rheumatic Causes of Positive ANA
100% Drug Induced Lupus
99% Lupus
97% Scleroderma
96% Sjogren’s
93% MCTD
80% Myositis
40% RA

32. ANA in Lupus Sensitivity 93-99% in SLE
Sensitivity % in drug induced Lupus
Specificity is not great
Higher the titre, higher the specificity
1:40- 30% normal population
1:160- seen in 5% of the population

33. Clinical Indications -ANA
ANA is NOT a good screening test given its low specificity
Presence of ANA does NOT mandate the presence of rheumatologic illness
A negative ANA is more useful and makes Lupus very unlikely
ANA titers correlate poorly with disease activity so serial measurements are not recommended
A positive ANA with anti-centromere pattern is very specific for limited scleroderma

34. Clinical Scenario: Case 2
42 year old caucasian female, otherwise healthy, comes to clinic complaining of fatigue. She complains of fatigue, poor sleep habits and aches and pains over the past year. Her joints have been bothering her, especially her hands. There is a discomfort and stiffness that comes and goes, and usually involves one hand at a time. She states that at times her hands have been mildly swollen and have limited her function.

35. Clinical Scenario Vitals T 99.2, BP 116/80, P 90 R 16 98%RA
PE is unremarkable
HEENT: WNL, no rash
CV: RRR, no murmurs
Pulm: CTA bilaterally
Abd: benign, no organomegaly
Ext: no edema, FROM of all joints, no appreciable joint swelling in wrist, MCP, PIP, DIP joints. No deformities. No rash

36. Clinical Scenario Lab Tests: CBC: WBC 6.3, Hg 12.7, Plt 266
Electrolytes: Na 138, K 4.2, Cr 0.7
TSH 3.8 (normal )
RF: negative
ANA: positive, titer 1:80
Anti TPO ab positive

37. Further testing of +ANA
This can be done to determine the exact nuclear target antigen
Some of these antibodies are specific for a particular disease
Include dsDNA, Smith, RO/SSA, La/SSB, U1RNP, Scl-70, centromere

38. Anti ds-DNA Specificity for SLE 97%
Present in about 60% of pt with SLE
Titers correlate with disease activity in SLE
Elevation correlates with Lupus nephritis
Seen in drug induced lupus

39. Anti- Smith Very specific for SLE >95%
See in only 20-30% of patients
No evidence that it is useful to follow for disease activity in SLE
Important diagnostic marker for SLE

40. Anti-Ro or SSA See in 70-97% of pt with Sjogren’s
See in 40% of SLE - associated with a photosensitive skin rash, lymphopenia, and Interstitial lung disease
In pregnant patients, associated with neonatal lupus and congenital heart block

41. Anti-La or SSB Usually see along with anti-Ro/SSA
Can see isolated activity in primary biliary cirrhosis and autoimmune hepatitis

42. Anti U1RNP A defining features for MCTD
Very sensitive for MCTD, but not specific, so use to rule out disease
Also found in 30-40% of pt with SLE

43. Anti-histone Antibodies
Seen in drug-induced lupus
Sensitivity of 100%
Not very specific, can see in 60-80% Lupus
Drugs commonly implicated:
hydralazine
INH
procainamide, penacillamine
quinidine

44. Anti Scl-70 Also known as anti-topoisomerase 1
Very specific for diffuse scleroderma
Specificity is greater than 95%
Sensitivity is low, range 22-40%
Higher levels associated with greater disease activity
Presence correlates with a higher risk of Interstitial Lung Disease

45. Anti-centromere ab
Usually associated with scleroderma, specifically CREST
Also see it in SLE, Raynaud’s
Sensitivity for Scleroderma ranges from 30-60%
Specificity for Scleroderma is high, greater than 95%

46. Clinical Scenario: Case 3
69 year old male at the VA, has known Hepatitis C. He comes into clinic complaining of generalized aches and pains in his joints. His left knee, right hand and right shoulder have been bothering him for a couple of months, and in the morning are stiff for 15 minutes. A Rheumatoid Factor is checked and this returns positive.

47. Rheumatoid Factor (RF)
What is it?
Autoantibody directed against the Fc portion of IgG, can be IgM or IgA

48. RF positivity
In what disease states do you see it?

49. RF positive disease states
Rheumatic Conditions
Infections
Pulmonary Disease
Misc RA
SBE
Silicosis
Aging
SLE MCTD
TB Leprosy
Sarcoidosis
Leukemia
Sjogren’s Syndrome
Syphilis
IPF
Colon Cancer
Systemic Sclerosis
Viral infections
Asbestosis
Cirrhosis- Hep C/PBC
Cryoglobulinemia
Parasitic Disease

50. Nonrheumatic RF+ Diseases

51. RF Positivity and Aging
Frequency of a positive RF increases with age
Age 20-60: 2-4%
Age 60-70: 5%
Age>70: %

52. Rheumatoid Arthritis

53. Rheumatoid Factor in RA
Sensitivity for RA: 80%
Note that up to 40% of patients with RA may be seronegative early on
Specificity for RA: 80-95%
Higher the titer or value of RF, higher the specificity for RA

54. Clinical Indications for RF
Little value as a screening test for RA
A positive RF does NOT equate with RA
In those patients with RA, a +RF usually predicts more aggressive erosive disease
Higher RF titers = higher specificity = higher positive predictive value for RA
Serial measurements are not indicated, and do not correspond with disease activity

55. Clinical Scenario
69 year old male at the VA, has known Hepatitis C. Comes into clinic complaining of generalized aches and pains in his joints. His left knee, right hand and right shoulder have been bothering him for a couple of months, and in the morning is stiff for 15 minutes. A Rheumatoid Factor is checked and this returns positive.
Anti CCP is negative.

56. Anti- CCP What are they? Antibodies to cyclic citrullinated peptide
Antibodies that target citrullinated proteins
Citrulline = a modified arginine amino acid
May be one of the major autoantigens driving the local immune response

57. Anti- CCP Sensitivity 50-75% Specificity greater than 90-95%
Found in low frequency in other rheumatic diseases
May be detected in patients with early RA
May predate the clinical development of RA by several years
Predictor of more erosive disease

58. Anti- CCP- Indications for Clinical Use
A disease-specific autoantibody that is very useful for the diagnosis of RA
Just as sensitive, and even more specific than RF
May predict eventual development of RA when found in undifferentiated arthritis
A marker of erosive disease

59. ANCAs What are they??? Anti-neutrophil cytoplasmic antibodies
How are they measured?
Two step procedure

60. ANCAs Step 1- Indirect Immunofluorescence Assay
Take patient serum and add it cells
If there are antibodies they will bind
Add a fluorochrome tag
View under a fluorescent microscope
If it lights up in the cytoplasm, then it is Cytoplasmic-ANCA (cANCA) positive
If is lights up around the nucleus, then it is Perinuclear-ANCA (pANCA) positive
Sensitive but not specific

61. Cytoplasmic-ANCA

62. Perinuclear-ANCA

63. ANCAs Step 2- Enzyme Immunoassay
Helps determine the specific antigen that the antibody is binding to
Two most common are Proteinase 3 (PR3) and Myeloperoxidase (MPO)
Not observer dependent
High specificity
High positive predictive value

64. Cytoplasmic-ANCA More specific for vasculitis
c-ANCA is associated with proteinase 3 (PR3)
Sensitivity reaches 90% in active generalized Wegener’s
Thus absence of ANCA does not rule out Wegener’s

65. p-ANCA Disease States Microscopic Polyangiitis Churg Strauss Syndrome
Pauciimmune Glomerulonephritis
Goodpasteur’s
Drug-Induced Vasculitis
Ulcerative Colitis
Crohn’s Colitis
Primary Sclerosing Cholangitis
Endocarditis
Malaria

66. Perinuclear-ANCA Less specific for vasculitis
It is associated with Myeloperoxidase (MPO)
Helpful in differentiating polyarteritis nodosa from microscopic polyangiitis

67. Clinical Indications for ANCA testing
Do not use it as a screening test
Using the pr3 and MPO increases the positive predictive value
Controversy regarding following ANCAs to monitor disease activity

68. HLA-B27 Human Leukocyte Antigen B-27
95% sensitivity for Ankylosing Spondylitis
80% sensitivity for Reactive Arthritis
Low specificity
Background prevalence of 6-10% in caucasian populations

69. Rheumatologic Testing
These labs are NOT useful as screening test
A positive test may or may not be associated with the disease
Selective ordering in patient with a high pretest probability
Ordering “Rheum Panel” is not recommended

70. From Cleveland Clinic
The diagnosis of rheumatologic diseases is based on clinical information, blood and imaging tests, and in some cases on histology. Blood tests are useful in confirming clinically suspected diagnosis and monitoring the disease activity. The tests should be used as adjuncts to a comprehensive history and physical examination.

71. What labs would you order?
1. A patient with known lupus admitted for flare of lupus nephritis

72. What labs would you order?
2. A patient with inflammatory arthritis involving PIPs, MCPs, wrists, knees admitted for a flare

73. Differential Diagnosis?
3. 58 year old patient with fevers and an ESR 99 and CRP of 10mg/dl