Clinical applications of Synovial (joint ) fluid analysis Dr.Sandeep C Agrawal Agrasen Hospital Gondia India Osteoarthritis,Rheumatoid Arthritis,Joint Tuberculosis,Crystal Induced Arthropathy,Periarticula Malignancy,SLE,Autoimmune diseases

1. Synovial Fluid ( Joint
Fluid ) Analysis
Dr.Sandeep Agrawal
Consultant Orthopedic Surgeon
MS,DNB
Agrasen Hospital
Gondia
Maharashtra
India
www.agrasenortho.com
drsandeep123@gmail.com
09960122234

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Synovial Fluid
 Synovial
 syn(like) + ovia (egg)
 “Joint Fluid”

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GENERATION AND CLEARANCE OF SYNOVIAL
FLUID
• The Starling equation reads as follows:
• J = K ([Pc − Pi] − σ[πc − πi])
• ([Pc − Pi] − σ[πc − πi]) is the net driving force,
• Kf is the proportionality constant, and
• Jv is the net fluid movement between
compartments.

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• According to Starling's equation, the
movement of fluid depends on six variables:
• Capillary hydrostatic pressure ( Pc )
• Interstitial hydrostatic pressure ( Pi )
• Capillary oncotic pressure ( πc )
• Interstitial oncotic pressure ( πi )
• Filtration coefficient ( Kf )
• Reflection coefficient ( σ )

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• Synovial fluid is a mixture of a protein-rich
ultrafiltrate of plasma and hyaluronan
synthesized by synoviocytes.
• Generation of this ultrafiltrate depends on the
difference between intracapillary and intra-
articular hydrostatic pressures and between
colloid osmotic pressures of capillary plasma
and synovial tissue fluid

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Synovial Fluid
 Viscous fluid found in the
cavities of movable joints
 Synovial membrane
 Inner membrane of
synovial joints
 Secretes synovial fluid into
the joint cavity
 Contain specialized cells
(synoviocytes)

7. Synovial fluid….Dialysate of plasma
Normal synovial fluid..clear pale or straw
coloured , viscous & does not clot
Contains 0.2 – 0.5% Hyluronan synthesised by
type-B synoviocytes
It polymerises to form large molecular weight
complexes causing high viscosity
Relatively acellular & has a WBC content of
<100x106/L :70% mono cytes; 30%
lymphocytes
SYNOVIAL FLUID

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• In normal joints, intra-articular pressures are
slightly subatmospheric at rest (0 to -5 )
• During exercise, hydrostatic pressure in the
normal joint may decrease further
• Resting intra-articular pressures in
rheumatoid joints are around 20 mm Hg,
whereas during isometric exercise, they may
increase to greater than 100 mm Hg, well
above capillary perfusion pressure and, at
times, above arterial pressure

9. Bulge test
The Bulge test is used to determine if there is an
abnormal amount of fluid surrounding a joint
Bulge test of joint for the detection of synovial effusion

10. Synovial fluid
Physical : viscosity,clarity,
colour.
Cytology: WBC count, smear for
Gram stain, AFB stain,
Bacteriology: culture &
sensitivity
Immunology: immune profile ,
R.A.
 Laboratory Testing
 Macroscopic Evaluation
 Chemical Examination
 Microscopic Examination

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Specimen Collection
 Arthrocentesis
Placement of needle in arthrocentesis of (A) elbow and
(B) knee joints.

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CollectionThree samples are collected.
Note
If the specimen cannot be examined immediately, fluid should be frozen and stored at -70°C until
examined

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Macroscopic Laboratory Testing
 Volume
 Color and Clarity
 Inclusions
 Viscosity
 Clotting
 Mucin Clot

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Macroscopic Analysis: Inclusions
 Rice bodies.
 Free-floating aggregates of tissue appear as rice
bodies.
 rheumatoid arthritis (RA)
 Degenarated synovium enriched with fibrin
 Ochronotic shards
 debris from joint prosthesis
 look like ground pepper
A =ochronotic shards
B =rice bodies

16. viscosity
Non- inflammatory conditions : Normal
vis.
Inflammatory : low viscosity due to
reduced content of hyaluronan & reduced
polymerisation (low molecular wt.)
String test: synovial fluid dripping from
pippette in the form of long string(10-
15cm)
Inflammatory : fluid falling as free
droplets

17. Colour & clarity
Deep yellow
In active R.A. slight greenish
tinge
Uniformly blood stained : PVNS
Loss of clarity : crystals,
increased cellularity, infective,
cartilage debris
Corporae oryzae ( rice
bodies):T.B., R.A.

19. WBC COUNT
T.C. INFLAMMATORY >2000 x
106/L
D.C.polymorphs, lymphocytosis,
synoviocytes, chondrocytes
Clumping of cells can be
prevented by collecting in EDTA
bottles, or heparin

20. Synovial fluid – table I
feature Non-inflam inflammato
ry
infective
example O.A. R.A. Septic
arthritis
viscosity high low low
colour Light straw Yellow-
greenish
Cream/
yellow
clarity clear Mild turbid opaque

21. Synovial fluid – table II
O.A. R.A./CRYST
ALS
SEPTIC
WBC
(x106/L)
50 - 500 1,500 –
30,000
>50,000
polymorphs occasional 30 – 70% >95%
smear normal Ragocytes,
macrophag
es,crystals
Polys,
bacteria
centrifugat
e
Cartilage &
fibrin
debris
Fibrindebris
/crystals
Fibrin
debris,
bacteria
R.A. -ve +ve -ve

23. eosinophils
Occasionally seen in normal
syn.fluid
Increased in haemarthrosis
Following arthrography –
contrast induced
Parasytic – Lyme arthritis

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Microscopic Analysis: Differential
 LE cells
 Neutrophils that have
engulfed a nucleus of a
lymphocyte
 Tart cells
 Monocytes that have
engulfed nuclear
material

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Microscopic Analysis: Differential
 Reiter cells
 Vacuolated macrophages
with ingested neutrophils
 RA cells
 “Ragocytes”
 Neutrophils with small,
dark, cytoplasmic granules
that consist of precipitated
rheumatoid factor

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Microscopic Analysis: Differential
 Hemosiderin
 Seen in Pigmented
Villonodular Synovitis
 Inclusions within clusters of
synovial cells
 Rice bodies
 Macroscopically resemble
polished rice
 Microscopically show
collagen and fibrin

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Laboratory Testing: Microbiology
 Staining
 Smears prepared by centrifugation or
cytocentrifugation
 Saline dilution reduces clustering of cells
 Gram’s stain most common
 Culture
 Set up with positive or negative stain results
 Aerobic
 anaerobic

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• Proteins are present in synovial fluid at
concentrations inversely proportional to
molecular size, with synovial fluid albumin
concentrations being about 45% of those in
plasma
• Synovial fluid is cleared through lymphatics in
the synovium, assisted by joint movement

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Chemical Analysis: Protein
 All proteins found in plasma
 Exception: various high–molecular weight proteins which
may be present in very small amount
 Fibrinogen
 beta 2 macroglobulin
 alpha 2 macroglobulin
 Use common serum protein procedures

30. Synovial fluid analysis
Recommended as a routine procedure in
the diagnosis of joint disorders by Ropes &
Bauer
Hollander introduced the term synovianalysis
Hollander & McCarty introduced polarised light
microscopy of synovial fluid for identification
of crystals of monosodium monourate &
calcium pyrophosphate
Biochemical analysis for glucose,protein,or
lactic acid are not useful

31. Synovial fluid analysis
Shmerling (1994) concluded :
Two major uses of synovial fluid
analysis were to identify or rule
out bacterial infection or
identification of crystals
A total of 6,556 papers were
published during 1980 – 2001 on
synovial fluid analysis

32. Infective arthritis
Difficult to differentiate infective
arthritis from acute crystal synovitis
on clinical examination alone in
monoarticular disease
Infection can co-exist with crystal induced
or rheumatoid inflammatory arthritis also
Synovial fluid analysis...Extremely useful to
identify crystals & bacteria and culture
studies

34. Crystal induced arthritis
Important cause of acute mono arthritis
Gout affects peripheral joints mostly and
pseudo-gout or CPPD mainly affects knee
joint
The converse is also true
Pain, swelling redness, sudden onset, and
fever mimic infective arthritis
Identification of crystals in syn.fluid is
diagnostic

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These fluid collections which serve as good samples of cloudy but translucent
inflammatory synovial fluid were taken from a patient with rheumatoid
arthritis (left) and gout (right) respectively

36. Traumatic arthritis
Following meniscal tears,
ligament injuries,
haemarthrosis, osteochondral
fractures, acute or episodic
synovitis of monoarticular
nature is common.
Syn. Fluid analysis along with
arthroscopy can establish the
diagnosis and help in treatment

37. Normal synovial fluid: Do not
clot
Clotting of synovial fluid =
fibrinogen
1.Damaged synovial
membrane
2.Traumatic tap
Macroscopic
Analysis: Clotting

38. Osteoarthritis
Acute on chronic monoarticular swelling is
quite common in degenerative arthritis
due to cartilage debris , avascular
necrosis, steroid usage,
haemoglobinopathies, sickle cell disease ,
alcoholism and diabetes
Tumour associated monoarthritis can
present without any evidence of
underlying pathology . SF analysis helps

39. Infective arthritis
In a large study(n=242) Weston
etal reported that SF Gram
staining was +ve in 50% of
cases only
SF culture was +ve in 67% of
cases
Blood culture was +ve
when SF gram staining & SF
culture were -ve

40. Syn.fluid analysis
Shmerling studied sensitivity and specificity of SF
analysis critically
He concluded that SF analysis was specific in
90% of infective arthritis and sensitivity of gram
staining was 50 – 75% and that of SF culture 75
– 95%
Freemont etal estimated that in patients on
antibiotic therapy SF cultures were +ve in 30 –
80% only and that usage of blood culture bottles
for SF culture and using large amount of Syn.fluid
improved pick-up rate

41. Infective arthritis
Freemont etal concluded that when
parameters like increased polymorph
cellcount >2000/cc and usage of special
culture media (BACTEC plus Anaerobic /F
medium) improved diagnostic sensitivity
In granulomatous lesions with little
aspiratable Syn.fluid, syn . tissue can be
used for culture purpose

42. Crystals in Syn.fluid
Absence of crystals in SF does not ruleout
crystal induced arthritis
Monosodium monourate crystals are
needle shaped, negatively bi-refrengent
and soluble in water
Crystals must be intra cellular
(phagocytosed) to diagnose gout
Extracellular cellular crystals in SF donot
produce gout

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 Monosodium urate (MSU)
 Calcium pyrophosphate (CPPD)
Crystal Identification

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 Corticosteroid
 Cholesterol
Crystal Identification

46. Intercritical gout
Pascual and Jovani studied 101
samples of syn.fluid from
asymptomatic gout
Results showed 43 out of 43
patients not on treatment for
gout had SF +ve for uric acid
crystals
34 out of 48 patients receiving
treatment for gout had uric acid
crystals in SF

47. Crystals in Syn.fluid
CPPD crystals : these are
associated with pseudogout
Rhomboid in shape and +vely
bi-refrengent
They also must be intra-cellular
to produce pseudo gout
Mere presence of these crystals
in SF is not diagnostic

48. Crystals in SF
Hydroxyapatite crystals are
found in syn. Fluid in Milwakee
arthritis
These cannot be seen under
polarising light microscopy
Only electron microscopy can
demonstrate these
crystals(TEM)
These crystals stain with Alizarin

49. Crystals in SF
Crystals of other substances like
steroids injected can be seen in
SF
Beclomethasone crystals are
needle shaped, but vary in size
and shape
Calcium oxalate crystals are
particularly seen in SF in renal
failure patients
Cholesterol crystals can be seen

50. Rheumatoid arthritis
Rheumatoid factor titres in
syn.fluid raise much before
seropositivity
In the presence of inflammatory
SF presence of RAF in SF is
diagnostic even if blood RAF is –
ve
RAF in SF is not only derived
from blood but also produced
by synoviocytes in joint

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Laboratory Testing: Rheumatoid Factor
 RF is an antibody to immunoglobulins.
 Present in rheumatoid arthritis:
 Serum – most cases
 Synovial fluid - 50%
 Rarely elevated only in synovial fluid and not serum
 False positives in other chronic inflammatory diseases.

52. Synovial fluid analysis
Mucin clot test: reflects de-polymarization
Of hyaluronic acid and can be
demonstrated by precipitation of
hyaluronate with acetic acid to form a clot
of mucin. One part of syn.fluid is added
to 4 parts of 2% acetic acid and briskly
stirred.
Its quality is interpreted

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Macroscopic Analysis: Viscosity
 “Ropes” or “Mucin Clot
Test”
 Normal = 4-6 cm
 When 2-5% acetic acid is
added, normal synovial
fluid will form a clot
surrounded by clear fluid

54. Mucin clot test
grade clot solution
good Tight ropy
mass
clear
fair Softer,shreddy Clear/hazy
poor shreddy turbid

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 “Ropes test”
 Estimation of hyaluronic acid–
protein complex integrity
 The adding of acetic acid to
normal synovial fluid, which
causes clot formation.
 Criteria:
 Compactness of the clot
 Clarity of the supernatant fluid
Macroscopic Analysis: Mucin Clot

58. String test
Syn.fluid is allowed to fall from
the pippette
Length of string is noted
Normal: 4 – 6 cm. String (5cm.
average)
Abnormal : less than 3 cm.
String indicates low viscosity or
inflammatory effusion

62. SF analysis vs Syn.biopsy
Johnson and Freemont (2000) published
a 10 year retrospective study of the
diagnostic usefulness of SF analysis and
syn. Biopsy
103 cases in which both SF analysis and
Syn.biopsy were performed were studied
Biopsy gave more information in 29% of
cases and SF analysis provided more
information in 18% of cases
In cases where syn. Biopsy was more
informative, than syn. Fluid analysis 65%
were inflammatory arthropathies and
35% were non-inflammatory

64. SF analysis vs syn.biopsy
In cases where SF analysis was
more informative, than syn.
Biopsy, 86% were inflammatory
and 14% were non-
inflammatory

65. Tuberculous arthritis
Wallace etal showed that
AFB +ve rate depended
upon the stage of disease.
20% of SF in Tuberculous
joints showed AFB +ve on
smear
80% of SF cultures for AFB
were positive in
tuberculous joints

66. Recent advances
Biochemical analysis of SF for glucose,
proteins and chloride is of little relevance
MMP3 levels in Syn.fluid reflect the
degree of inflammation of a joint and it
also correlates with TNF alpha which is a
potent inducer of of MMP-3 (matrix
metallo protease –3) in synovial
fibroblasts, but also of IL- 6 which in turn
increases levels of IL -1

67. Rheumatoid arthritis
IL-1 stimulates production of MMP3
MMP3 levels in SF correlate well with SF
IL1 levels aswell as with those of its
receptors sTNF-RI and sTNF-RII thus
corresponding to erosive rheumatoid
disease
conversely low SF levels of MMP3
correlate with non-erosive rheumatoid
disease
Detection of MMP3 thus indicates local
inflammatory reaction in the joint and it is
easy to detect because its concentration
is 1000times higher than that of IL-6

68. Rheumatoid arthritis
Measurement of IL6 remains difficult
because of its short half life, presence of
blocking factors , and its circadian rhythm
Though MMP3 is a synovium derived
indicator of inflammatory process, it can
also be measured in serum and correlate
well with levels of IL-6 and erosive
rheumatoid disease – positive predictive
value

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Classification of Joint Disorders

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71. 71

72. To have a fulfilling life, you should
find your life purpose and follow it
faithfully. Find what
matters to you and build the
courage to follow it. Having a one-
liner will help you
internalize and communicate with
your mission.
Happiness will never come to
those who fail to appreciate
what they already have.
- Unknown
The more passions and desires
one has, the more ways one
has of being happy.
-Charlotte-Catherine

73. 73
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