2. PRESENTATION LAYOUT
• Introduction to fluorescein
• Basic principle of fluorescence
• Some terminologies
• Anatomical considerations
• Indication and contraindication of FFA
• Procedure of FFA
• Normal phases of FFA
• FFA interpretation
• FFA Vs ICG
• Phenomenon of FFA
3. INTRODUCTION OF FLUORESCEIN
• Orange water soluble dye
• When injected IV , it remains largely intravascular and circulates in
blood stream
• 70 – 85 % of fluorescein bind with blood serum albumin
• Rest of remain free i.e unbound form
• Sodium fluorescein : C20 H10 O5 Na2
4. CONTINUE…
• Properties
-Non-expensive
-Non-toxic
-Flouresces at blood PH level 7.37 – 7.45
-Rapid diffusion
• Synthesized from the petroleum derivatives resorcinol and phthalic anhydride
Chemically related to Phenolphthalein
• Molecular weight : 376 daltons
5. BASIC PRINCIPLE OF FLUORESCENCE
• Absorption followed by release of the radiant energy in the form of
visible light
• Fluorescent substance follows Stokes law
Stokes law
• Fluorescent substances absorbs light
• Molecular excitation
• Electrons elevated to higher less stable state Returns to stable lower energy
form Releasing light of longer wavelength FLUORESCENCE
• Entire process – 10 – 8 sec.
6. BASIC PRINCIPLE OF FLUORESCENCE
• Absorption spectrum of fluorescein – 465 to 490 nm
• Excitation peak 490nm (blue part of spectrum )
• Emission spectrum of fluorescein – 520 to 530 nm
• Emission peak 530 nm (green-yellow spectrum )
7. NOTE
Absorbed radiant energy > emitted energy
AND
As energy – inversely proportional to –wavelength
SO,
λ of emitted wave > λ of absorbed wave
8. FILTERS FOR PROCEDURE
• Two type
• 1) blue barrier filter
• 2) yellow green barrier filter
• Blue barrier filter ensures that only the blue light enters the eye
9. • Yellow green barrier filter blocks the blue light reflects from the eye
• It allows green light to pass through unimpaired, to be recorded on the
film
10. TERMINOLOGIES
Fluorescence- ability of a compound to absorb light of shorter
wavelength and emit light of longer wavelength with in a very short
interval
Hyper-fluorescence – an area of abnormally high fluorescence due to
increase density of dye molecule
Hypo-fluorescence - an area of abnormally poor fluorescence
Auto-fluorescence – an inherent property of a lesion to spontaneously
fluoresce even in absence of dye
( observed before injection of the dye)
11. Arm retina circulation time- from dye injection to first
appearance in retinal arteries( 10-12 sec)
Pooling- accumulation of dye in closed space .e.g. RPE
detachment, CSR
Leakage- dye escapes in open space e.g. vitreous space
Window defect- type of early hyper-fluorescence due to RPE
atrophy
Control photograph –photo taken before dye given to detect
auto-fluorescence
12. Staining- late hyperfluorescence due to adsorption of the dye by a
tissue
Blocked fluorescence – hypofluorescence occurs by masking
underlying retinal and choroidal tissue by blood , pigment etc.
Capillary nonperfusion – due to non filling of the retinal capillaries
due to anatomical and function reasons
Artifacts- undesirable shadows that are seen following the
development of the film
13. ANATOMICAL CONSIDERATION
• Major choroidal vessels are impermeable to both bound and unbound
form of fluorescein
BUT
Choriocapillaries
• Walls are extremely thin
• Contains multiple fenestrations
• Through which free molecules pass across Bruchs membrane
Choriocapillaries and Bruchs membrane both permeable to free and
bound fluorescein molecules
14. CONTINUE….
• Outer blood retinal barrier : tight junction between RPE cells prevent
passage of fluorescein
• Inner blood retinal barrier : tight junctins between endothelium cells of
retinal blood vessels prevent passage of fluorescein
• Fluorescein pass from choriocapillaries also passes through bruch’s
membrane but it encounter with tight junction intracellular complex zonula
occludens of RPE cells and prevent passage
• Disruption of barrier leak both bound and free fluorescein molecules
15.
16. BLOOD VESSLES IN RETINA
• For FFA interpretation sensory retina divided into two layers
1. Inners vascular half ( ILM – INL )
Here retinal blood vessels located in two separate planes
large retinal arteries and veins located in nerve fiber layer
Retinal capillaries located in inner nuclear layer
2. Outer avascular half (OPL – RPE )
• when retina becomes edematous ,
• it is the layer that fluid accumulate causing the cystoid space
17.
18. PURPOSE OF FULUROSCEIN ANGIOGRAPHY
• Studying the normal physiology of the retinal and choroidal circulation,as
well as disease process affecting the macula.
• Evaluation of the vascular integrity of the retinal and choroidal vessels
• Check the integrity of the blood ocular barrier.
- Outer blood retinal barrier breaks in CSR
- Inner blood retinal barrier breaks in NVD, NVE
19. CONTINUE…
• It helps in clinical diagnosis
• To determine extent of damage
• To formulate treatment strategy for choroidal and retinal disease
• To monitor result of treatment
20. INDICATION OF FFA
Retinal vascular malformation and tumors
Retinal vascular disorders
Macular disorders
Choroidal disorders
Optic nerve disorders
24. CONTRAINDICATIONS
ABSOLUTE
1) known allergy to iodine containing compounds.
2) H/O adverse reaction to FFA in the past.
RELATIVE
1) Asthma
2) Hay fever
3) Renal failure
4) Hepatic failure
5) Pregnancy ( especially 1st trimester)
25. MILD MODERATE SEVERE
Staining of skin,
sclera and mucous
membrane
Nausea and
vomiting
Respiratory-
laryngeal edema
,bhroncospasm
Stained secretion
Tear, saliva
Vasovagal response Circulatory shock,
MI, cardiac arrest
Vision tinged with
yellow
utricaria Generalized
convulsion
Orange-yellow urine fainting Skin necrosis
Skin flushing,
tingling lips pruritis
periphlebitis
COMPLICATIONS
26. COMPICATIONS MANAGEMENT
• Unavoidable minor side effects : treatment not needed
• Temporary tan skin colour, Red after image from the photoflash and
discoloration of the urine
• Transient Nausea and vomiting (10%): treatment not needed
• Vasovagal syncope (1%) :treatment not needed
• In extreme bradycardia
• IV atropine may be needed.
27. CONTINUE..
• Anaphylaxis such as bronchospasm, urticarial skin rash and
hypotension (<1%).
• Treatment is with chlorpheniramine (piriton) 10mg IV, hydrocortisone
100mg IV
• Hypotension and Bronchospasm
• oxygen and adrenaline 1ml of 1:1000 IM
• Cardiac and respiratory arrest (<0.01%)
• Treatment would involve cardiopulmonary resuscitation
28. EQUIPMENT AND MATERIALS NEEDED FOR ANGIOGRAPHY
Fundus camera and auxilliary equipment
Matched fluorescein filters ( barrier and exciter )
Digital photoprocessing unit ( computer based )
23 gauge scalp vein needle
5 ml syringe
5 ml of 10% OR 3ml of 25 % fluorescein solution
29. 20 gauge , 1.5 inch needle to draw the dye
Armrest for fluorescein injection
Tourniquet
Alcohol swabs
Bandage
Standard emergency equipment
30. PROCEDURE
Patient is informed of the normal procedures, the side effects and the adverse
reactions.
Dilating the pupil
Made to sit comfortable.
3-4 red free photographs taken.
(control photographs)
5ml of 10% or 3ml of 25% NAF injected through the anticubital vein
31. Wait for 8 seconds for young and 12 seconds for older patients
( normal arm-retina time)
Photos are taken at 1 second interval for 10 seconds
Then every 2 seconds interval for 30 seconds
Late photographs are usually taken after 3 ,5 and 10 minutes.
32. CIRCULATION OF DYE
Dye injected from peripheral vein
venous circulation
heart
arterial system
INTERNAL CAROTID ARTERY
Ophthalmic artery
Short posterior ciliary artery) Central retinal
(choroidal circulation.) ( retinal circulation)
33. NORMAL PHASES IN FFA
•Early phase
• Choroidal(prearterial)
• Arterial
• Arteriovenous (capillary)
• Venous
• Early
• mid
• Late
•Mid phase
•Late phase
34.
35. CONTINUE…
• Normally 10 -15 secs elapse between dye injection and arrival of dye
in the short ciliary arteries
• Choridal circulation preceeds retinal circulation by 1 Sec
• Transit- if dye through the retinal circulation takes approximately 15-
20 secs
36. EARLY PHASE
• Choroidal filling through the short ciliary arteries
• Initial patchy filling of lobules followed by diffused blush
as dye leaks out of choriocapillaries
• Cilioretinal vessels and prelaminar vessels and
prelaminar optic disc capilaries fill
Choroidal ( prearterial ) phase
37.
38. FACTS OF PATCHY CHOROIDAL FILLING
• Choriocapillaries has number of lobules
• The lobules fill independently from one another,
• giving a transiently patched or blotched appearance
39. ARTERIAL PHASE
• Begins with the first appearance of fluorescein in the arteries, and
extends until the arteries are completely filled
• Posterior pole fills with dye earlier than the periphery
• Superior branches usually fill first
41. ARTERIO-VENOUS PHASE(CAPILLARY PHASE)
• Complete filling of retinal arteries and capillaries.
• Early laminar flow in the veins in which dye is seen along the
lateral wall of the vein
• Choroidal fluorescence increases as free fluorescein continues to
leak from the choriocapillaries
43. VENOUS PHASE
• Gradually whole diameter of the veins is filled
• Earliest seen in the peripapillary and macular region
• Divided according to the venous filling and arterial emptying
• Early
• mid
• Late
44. EARLY VENOUS PHASE
• Arteries and capillaries are completely filled and marked lamellar
venous flow
45. MID VENOUS PHASE
• Some veins are completely filled
• Some shows marked laminar flow
46. LATE VENOUS PHASE
• All veins are completely filled and the arteries beginning to empty
47. MID PHASE
• Known as recirculation phase
• 2-4 min after injection
• Veins and arteries remain roughly equal in brightness.
• Intensity of fluorescein diminishes slowly as
• flourescein is removed from the blood stream on the first pass through
the kidneys.
48. LATE PHASE
• After 10-15 minutes little dye remains in the blood stream
• This phase demonstrates
• Gradual elimination of the dye from the retinal and choroidal
vasculature
• staining of optic disc , sclera is normal finding
• Any other hyperfluoresecence suggest the presence of abnormality
50. Phases of angiogram Time ( in seconds)
Injection 0
Posterior ciliary artery 9.5
Choroidal phase 10
Arterial 10 - 12
Arterio venous 13
Early venous 14 - 15
Mid venous 16 -17
Late venous 18 – 20
Late ( elimination) 5 minutes
51. FLUORESCENCE IN FOVEAL REGION
• Dark appearance WHY?
i) Avascularity in the FAZ
ii) Blockage of the choroidal
flourescein because of
• increased amount of xanthophyll
pigments at fovea
• melanin in RPE
52. NORMAL ANGIOGRAM
• Patchy filling of choroid
• Retinal blood vessels filling
• Dark area of foveal avascular zone
• But there is no hyper or hypofluroscence area
• At the end of the transit phase, fluorescein dye remains in the choroid
and sclera due to leakage from the choroidal vessels
• A small amount of fluorescein also remains in the optic nerve head and
retinal vessels, but there is no leakage
• Any additional fluorescein in the eye should be regarded as pathologic
54. STEPWISE APPROCH TO FFA
• A fluorescein angiogram should be interpreted systematically to
optimize diagnostic accuracy as follows:-
• A ) Indicate whether images of right , left or both eyes have been
taken.
• B)comment on the red free images
• C)indicate any delay in filling as well as hyper or hypo
fluorescence
• D)indicate any characteristic features such as a smoke –stack or
lacy filling pattern.
54
55. FFA INTERPRETATION FLOW CHART
Fluorescein angiogram
Normal Abnormal Auto/pseudofluorescence
Hyperfluorescence Hypofluorescence
Leakage Pooling Staining Window Blocked Non
defect filling
56. NOTE
• Hyperfluorescence and hypofluorescence can alternate in same
location
• Especially in inflammatory disorder
• 1st hypofluorescence due to retinal oedema
• Later hyperfluorescence due to increased vascular permeability
57.
58. AUTOFLUORESENCE
• Emission of fluorescence light in the absent of fluorescein
Example : optic nerve head drusen , astrocytic hematoma ,
myelinated nerve fibers
Optic disc drusen Astrocytic hematoma
59. PSEUDOFLUORESCENCE
• Occurs when nonfluorescence light passes through the entire filter system
• Blue reflected light passes from green filter pseudofluorescence occurs
• It decrease contrast aswell as resolution of image
• To avoid pseudofluorescence filter combination to be sure that no significant
overlap exists
• Over the time filter alter the range of light transmission so should be change
in certain time . Auther recommend about 5 year time to change filter
60. WINDOW DEFECT
• Focal RPE atrophy
• Unmasking of normal background of choroidal fluorescence
• Characterized by early hyperfluorescence which increases in intensity
then fade without changing shape and size
e.g. inflammation of RPE
atrophy of RPE , drusen
61.
62. EXTRAVASCULAR LEAK
• Pooling and staining in choroid
• Cystoid edema and noncystoid edema in retina
• Neovascularization , inflammation and tumor vessels in vitreous
• Disc staining
Cystoid oedema of macula
63. Pooling( accumulation of dye
in a closed space)
-Early hyperfluorescence
sub-retinal space Early hyperfluorescence
sub RPE space
increase in size ,intensity increase intensity only
e.g. CSR e.g. PED
67. STAINING
• Accumulation of fluorescence within a tissue
• Due to prolonged dye retention
• Minimum hyperfluorescence in early and midphase which increases
in late phase
• Can be seen in normal as well as pathologically altered tissue
74. BLOCKED FLUORESCENCE
• Optical obstruction (masking) of normal density of fluorescein
• Caused by lesions anterior to retina
• Pre-retinal lesions eg.vitreous opacity,preretinal
haemorrhage block all fluorescence
• Deep retinal lesions eg.intraretinal haemorrhage and
hard exudates block only capillary fluorescence
• Increased density of RPE eg.congenital hypertrophy
• Choroidal lesions eg.naevus
EXAMPLE
75.
76.
77.
78.
79. FILING DEFECTS
• Inadequate perfusion of tissue with resultant low fluorescein content
• Avascular occlusion of choroidal circulation or retinal
arteries,veins and capillaries
• Loss of vascular bed eg.severe myopic degeneration –
choroideremia
• Emboli
• arteriosclerosis
EXAMPLE
81. LIMITATIONS OF FFA
1) Does not permit study of choroidal circulation details
due to
a) melanin in RPE
b) low mol. Wt. of fluorescein
how to overcome ---- ICG
2) More adverse reaction
3) Inability to obtain angiogram in patient with excess
hemoglobin or serum protein
82. INDOCAINE GREEN ANGIOGRAPHY
• FFA excellent method for demonstrating retinal
circulation.
• But…
• Not helpful in delineating choroidal circulation
• ICG –of particular value in studying choroidal
circulation ,
• Can be useful adjunct to FA in investigation of macular
diseases.
83. FFA Vs ICG
PARAMETERS FFA ICG
1) Dye used Sodium fluorescein Indocyanine
green
2) Light used visible spectrum infrared
3) purpose study retinal
vasculature
Choroidal
vasculature
4) Filter used Blue- green Infra-red
5) expense lower higher
84. PHENOMENON OF FFA
• All the process of occurrence of hyper or hypo-fluorescence can be
described under following 3 phenomenons
A. OPTICAL PHENOMENON
B .MECHANICAL PHENOMENON
C. DYNAMIC PHENOMENON
85. OPTICAL PHENOMENON
• Normal neurosensory retina is transparent
• Normal RPE and Bruch’s Membrane are semitransparent
• Hence, we can see choroidal fluorescence
• BUT, this transparency can be pathologically increased or decreased
86. DECRESEING TRANSPARENCY
• In case of blocked fluorescence ,
transparency is lost
• SO, WE DO NOT SEE CHOROIDAL FLUORESCENCE
91. MECHANICAL PHENOMENON
• Related to adhesion of RPE to Bruch’s Membrane
• RPE firmly attached to Bruch’s membrane by hemidesmosomes
92.
93. Absence of hemidesmosomes
RPE splits away from Bruch’s membrane
Fluorescein stained fluid accumulate in between them
eg. PIGMENT EPITHELIAL DETACHMENT
94.
95. DYNAMIC PHENOMENON
• Related to diffusion of fluorescein in ocular tissue
• Determined by inner and outer blood retinal barrier I.E DIFFUSION
BARRIER
96. RETINAL VESSELS
• Normal retinal vessels do not leak fluorescein - due to zonula
occludents in between endothelial cells
• These zonula occludents open up during inflammatory process
101. RETINAL PIGMENT EPITHELIUM
• Normal RPE is tight
• zonula occludens seal portion of all the intercellular spaces of the
pigment epithelial monolayer.