1. Tutor Hematologi Putaran I
PEMERIKSAAN
STATUS BESI TUBUH
Lulut Kusumawati, dr / Fery H Soedewo, dr, MS, SpPK(K)
1
2. Pendahuluan
• Besi dalam tubuh berguna untuk :
pembentukan
hemoglobin, mioglobin, berbagai enzim
• Kandungan normal besi tubuh :
• Laki-laki : 50 mg/kgBB
• Wanita : 35 mg/kgBB
• Dalam tubuh besi selalu berikatan dengan
protein tertentu
2
3. Absorbsi Besi
• Tergantung : kebutuhan tubuh – rasio
enhancer inhibitor
• Di duodenum proksimal dan jejunum
• Sebagian besar diet dalam bentuk non
heme (Fe3+)
• Enhancer : askorbat, sitrat, daging
• Inhibitor : phytat, tannin
• Fe3+ Fe2+ (supaya bisa diserap)
• Jumlah : 1 mg yang masuk darah
• Dalam plasma besi dalam bentuk Fe3+
• terikat transferrin 3
4. Distribusi dan transportasi
• Protein yang berperan dalam transportasi
dan penyimpanan besi :
o Apotransferrin
o Apoferritin
o Reseptor transferrin
• Besi disimpan dalam bentuk ferritin dan
hemosiderin
4
5. Transferrin
SI Unsaturated Iron Binding
Capacity (UIBC)
SI + UIBC =
( TIBC )
5
8. Persiapan Tes
• Pasien berhenti obat oral Fe 12 jam
Sampel tes sebelumnya
• Serum atau plasma heparin
• Tabung plastik disposable 12x75 mm
Peralatan • Glassware : cuci deterjen – rendam HCl
2 mol/L (6-12 jam) – bilas iron free water
Iron free • Deionized water
• 0,2 µmol besi/liter
water
8
9. Serum Iron (di PK)
Prinsip :
• Ikatan antara ferri (Fe3+) dan transferrin
dilepaskan oleh guanidine dalam suasana
pH 4,8
• Asam askorbat akan mereduksi ion ferri
(Fe3+) menjadi ferro (Fe2+)
• Fe2+ akan bereaksi dengan ferrozine
membentuk komplek berwarna.
• Dibaca dengan fotometer pada panjang
gelombang 560 nm.
9
10. Serum Iron (di PK)
Reagen :
• Reagen 1 (R1) :
o Acetate buffer, pH 4,8 100 mmol/L
o Guanidine hydrochloride 5 mol/L
o Thiourea 52,5 mmol/L
• Reagen 2 (R2) :
o Ascorbic acid
• Reagen 3 (R3) :
o Ferrozine 41 mmol/L
• Standar : Std
o Iron 100 μg/dL
1 mg/L
17,9 μmol/L
Sampel : serum atau plasma heparin. Sampel tidak
boleh hemolisis 10
11. Serum Iron (di PK)
1. Membuat larutan kerja (working
reagent) :
o Larutkan 1 sendok takar R2 dalam 50 ml R1.
Tunggu hingga tercampur dengan baik.
1 sendok
takar R2
50 ml R1
o Stabil selama 2 minggu pada suhu 2-8˚C dan 3
hari pada suhu 20-25˚C.
11
12. Serum Iron (di PK)
• 2 BLANKO STANDAR SAMPEL
Akuades Standar Sampel
150 μL 150 μL 150 μL
Working Working Working
reagent reagent reagent
500 μL 500 μL 500 μL
A1 A2
Tunggu 50 detik, kemudian baca OD pada panjang
gelombang 560 nm (A1 dan A2)
12
13. Serum Iron (di PK)
• 3.
R3 25 μL R3 25 μL R3 25 μL
A3 A4
Baca OD setelah 325 detik, pada panjang
gelombang 560 nm (A3 dan A4)
13
14. Serum Iron (di PK)
• Penghitungan :
A4-A2 x konsentrasi standar (n)
A3-A1
µg/dL n = 100
mg/dL n = 1
µmol/L n = 17,9
• Nilai rujukan : 0,5 – 1,68 mg/dL
50 – 168 µg/dL
8,95 – 30 µmol/L
14
15. PK ICSH
Guanidine Protein Trichloracetic
hydrochloride precipitant acid dan
thioglycollic
acid
17,9 μmol/L Larutan besi 80 μmol/L
standar
560 nm Λ fotometer 562 nm
Tanpa Sentrifus 13.000 g, 4
sentrifugasi menit
• 50 detik Waktu • 300 detik
• 325 detik tunggu • 600 detik
reaksi 15
16. Total Iron Binding Capacity (TIBC) di PK
Metode : Saturasi
Prinsip :
TIBC dievaluasi setelah transferrin sampai
jenuh oleh larutan besi, dan kelebihan
besi akan diabsorbsi oleh magnesium
hydroxide carbonate. Setelah
disentrifus, konsentrasi besi dalam
supernatan diukur.
16
17. Total Iron Binding Capacity (TIBC) di PK
Reagen :
• Reagen 1 : R1
o Iron saturating solution 500 μg/dL
5 mg/L
89,5 μmol/L
• Reagen 2 : R2
o Magnesium hydroxide carbonate (1 sendok
takar = 100 mg)
Sampel : Serum, plasma heparin. Sampel tidak boleh
hemolisis
17
18. Total Iron Binding Capacity (TIBC) di PK
1 sendok
1 ml
takar reagen
reagen 5 menit R2 Inkubasi 20
R1
menit, sekali-kali
dikocok
0,5 ml
sampel
Sentrifus 3000
rpm, 10 menit
Periksa supernatan seperti pada SI Ambil supernatan
Dengan supernatan : reagen = 1:3
18
19. Total Iron Binding Capacity (TIBC) di PK
Nilai rujukan :
o Laki-laki dewasa : 2,6 - 3,9 mg/L
= 46,5 – 69,8 μmol/L
= 260 -390 μg/dL
o Wanita dewasa : 2,1 – 3,4 mg/L
= 37,6 – 60,9 μmol/L
= 210 – 340 μg/dL
19
22. Interpretasi SI dan TIBC
SI :
o Normal fluktuasi lebar dan memperlihatkan
variasi diurnal
o Tidak terpengaruh sampai cadangan besi
habis.
o Penurunan SI terjadi pada anemia defisiensi
besi, penyakit kronis dan selama respon fase
akut
TIBC :
o TIBC meningkat pada anemia defisiensi besi
dan kehamilan dan menurun pada infeksi dan
keganasan.
22
23. Ferritin Serum
Metode : ELISA metode double sandwich
Prinsip :
Antibodi dengan high affinity terhadap
ferritin (antiferritin Ig G) akan berikatan
dengan feritin serum dan selanjutnya
dilabel dengan enzim horseradish
peroxidase dan dibaca OD-nya pada
panjang gelombang 492 nm.
23
24. Ferritin Serum
Metode : IRMA (Immunoradiometric Assay)
Prinsip :
Antibodi yang dilabel dengan
radioaktif yang berlebih direaksikan
dengan ferritin. Ferritin yang tidak
berikatan dengan antibodi akan
dihilangkan dengan
immunoadsorbent.
24
27. Hemosiderin
Prinsip :
reagen Prussian blue akan mewarnai besi
menjadi berwarna biru terang atau hijau, eritrosit
tercat warna merah atau merah muda oleh
neutral red.
Bahan dan alat :
• Gelas obyek dan bahan sampel hapusan
darah
• Metanol untuk fiksasi
• Potassium ferrocyanide 10 g/l dalam 0,1 mol/l
HCl
• Larutan neutral red atau eosin
• HCl 3 mol/l
27
28. Hemosiderin
Hapusan darah difiksasi dengan
metanol, tunggu 10 – 20 menit
Masukkan dalam larutan
potassium ferrocyanide (10 menit
- 200C)
Cuci dengan air kran (20
menit), bilas dengan distilled
water.
Counterstain dengan neutral red
atau eosin selama 10-15 detik
Tuangi dengan HCl 3 mol/l dan
cuci dengan air kran.
28
29. Prussian-blue staining (Pearl’s reaction) on aspirated bone
marrow particles to demonstrate iron stores.
A. Normal B. Absent C. Increased D. Grossly increased
29
31. Enhancer - Inhibitor
• Enhancer : bahan-bahan yang meningkatkan
absorbsi
Askorbat (mereduksi Fe3+ Fe2+, sitrat, asam
organik lain dalam sayuran
• Inhibitor : bahan-bahan yang menghambat
absorbsi
Karbonat, phytat, tannate, phosphat, oksalat
chelating agent untuk Fe
31
32. • Meningkatkan absorbsi :
Daging, ASI, cairan lambung
• Menghambat absorbsi :
Putih telur, protein susu sapi, cairan pankreas
32
34. • Plasma-EDTA :
warna yang terbentuk lebih lambat dan harus
didiamkan sekitar 15 menit sebelum dibaca
dengan fotometer
• Sampel tidak boleh hemolisis :
Hemolisis akan melepaskan besi dalam plasma
sehingga mengganggu hasil pemeriksaan
34
35. Deionized Water
• Also known as demineralized water[
• Water that has had its mineral ions removed,
cations : sodium, calcium, iron, copper
anions : chloride and bromide.
• Deionization is a physical process which uses
specially-manufactured ion exchange resins
which bind to and filter out the mineral salts from
water.
• Deionization does not significantly remove
uncharged organic molecules, viruses or
bacteria, except by incidental trapping in the
resin. Specially made strong base anion resins can
remove Gram-negative bacteria. Deionization
can be done continuously and inexpensively using
electrodeionization.
35
36. HCl = Hydrochloric Acid
• Hydrogen chloride (HCl) is a monoprotic acid, which
means it can dissociate (i.e., ionize) only once to give
up one H+ ion (a single proton). In aqueous
hydrochloric acid, the H+ joins a water molecule to
form a hydronium ion, H3O+
• HCl + H2O → H3O+ + Cl− The other ion formed is
Cl−, the chloride ion. Hydrochloric acid can therefore
be used to prepare salts called chlorides, such as
sodium chloride. Hydrochloric acid is a strong
acid, since it is essentially completely dissociated in
water.
36
37. HCl = Hydrochloric Acid
• One of the most important applications of
hydrochloric acid is in the pickling of steel, to remove
rust or iron oxide scale from iron or steel before
subsequent processing, such as
extrusion, rolling, galvanizing, and other techniques.
• Fe2O3 + Fe + 6 HCl → 3 FeCl2 + 3 H2O
The spent acid has long been re-used as iron(II)
chloride (also known as ferrous chloride) solutions, but
high heavy-metal levels in the pickling liquor have
decreased this practice.
• 4 FeCl2 + 4 H2O + O2 → 8 HCl+ 2 Fe2O3
By recuperation of the spent acid, a closed acid loop
is established. The iron(III) oxide by-product of the
regeneration process is valuable, used in a variety of
secondary industries. 37
38. Kandungan Besi Tubuh
• Plasma pool : SI – TIBC – saturasi transferin
• Storage pool : Feritin - Hemosiderin
• Red blood cell pool : reseptor transferin –
protoporfirin eritrosit – indeks eritrosit – RDW –
hemoglobin – hematokrit.
38
39. Larutan Besi Standar 80
µmol/l
• 22,1 ml deionized water + 200 µl 2 mol/l HCl
• Campur
• + 100 µl larutan besi standar (1000 µg Fe/ml dalam 1%
HCl, Aldrich No 30, 595-2)
• Simpan dalam suhu kamar selama 2 bulan.
39
40. Larutan kromogen
• 25 mg ferrozine + 100 ml 1,5 mol/l sodium acetate
• Simpan dalam keadaan gelap suhu kamar selama
1 bulan
40
41. Serum Iron (ICSH)
Prinsip :
• Besi dilepaskan dari ikatannya dengan
transferrin.
• Direduksi dari Fe3+ menjadi Fe2+ dan serum
protein
• Presipitasi
• Fe2+ + larutan kromogen komplek warna
(pink solution)
• Dibaca dengan fotometer pada panjang
gelombang 535 nm.
41
42. Serum Iron (ICSH)
Reagen :
• Protein precipitant
100 g/L trichloracetic acid (0,61 M) dan 30
ml/L thioglycollic acid dalam 1 mol/L HCl.
• Larutan kromogen
25 mg ferrozine dilarutkan dalam 100 ml
sodium acetate 1,5 mol/L
• Larutan standar besi (80 μmol/l)
42
43. Serum Iron (ICSH)
0,5 ml 0,5 ml 0,5 ml
protein protein protein
precipitan precipitan precipitan
0,5 ml serum 0,5 ml larutan 0,5 ml iron-free
standar water
Tunggu 5 menit
Sentrifugasi 13000 g Jika tak ada Microfuge :
selama 4 menit sentrifugasi 1500 g
supernatan (Microfuge) selama 15 menit dgn
volume dobel
43
44. Serum Iron (ICSH)
0,5 ml 0,5 ml
0,5 ml
larutan larutan
larutan
kromogen kromogen
kromogen
0,5 ml larutan 0,5 ml iron-free
0,5 ml supernatan standar + water + protein
protein presipitant
presipitant
Tunggu 10 menit
Baca absorbans pada panjang gelombang 562 nm
44
45. Serum Iron
• Penghitungan :
Ates-Ablanko x konsentrasi standar
Astandar-Ablanko
• Nilai rujukan :
o Anak-anak : 0,5 – 1,2 mg/L
( 9,0 – 21,5 μmol/L)
o Wanita dewasa : 0,5 – 1,7 mg/L
( 9,0 – 30,4 μmol/L)
o Laki-laki dewasa : 0,65 – 1,75 mg/L
( 11,6 – 31,3 μmol/L)
45
46. Total Iron Binding Capacity (ICSH)
• Dalam plasma, besi terikat pada
tranferin, dan TIBC adalah mengukur
protein tersebut.
• Prinsip :
Serum ditambahkan kelebihan besi
(Ferri klorid). Besi yang tidak terikat
transferin akan diabsorbsi oleh
magnesium carbonate, kemudian
kadar besi serum diukur.
46
47. Total Iron Binding Capacity (ICSH)
Reagen :
• Basic magnesium carbonate
• Saturating solution (100 μmol Fe/L).
• 17,7 ml deionized water + 100 μL HCl +
1 mol/L 100 μL larutan standar.
(Saturating iron solution mengandung
5,6 μg Fe/mL)
47
48. Total Iron Binding Capacity (ICSH)
0,5 ml 100 mg
Campur dan magnesiu
saturatin
diamkan 15 m
g iron
menit pada carbonat
solution
suhu ruangan
kocok
0,5 ml serum
Diamkan selama 30 menit,
sekali-kali dikocok
Periksa Sentrifugasi, 13.000 g selama
seperti SI Ambil 0,5 ml supernatan
4 menit
48
49. Ferritin Serum - IRMA
• This a highly sensitive test.
• It uses radioisotopes as labels to detect the presence
of antigen (ferritin) or antibody in a sample.
• Unlabeled ("cold") antigens compete for the
antibodies.
• Radioactive antigens are displaced from the
antibodies.
• The antibody-bound antigen is separated from the
free antigen in the supernatant fluid.
• The radioactivity of each bound antigen is measured
with a scintillation counter.
49
51. Ferritin Serum - ELISA
• The patient’s serum containing the antigen
(ferritin) is added in Microwell strips (which
contains the antibody).
• A second antibody coupled to an enzyme is
then added (forming a "sandwich").
• The substrate (chromogen) solution is added
and will be cleaved by the enzyme to form a
colored product (qualitative assessment).
• This can be measured quantitatively by a
spectrophotometer.
51
53. Feritin Serum
Reagen :
• Preparat antiferitin Ig G yang dikonjugasi
dengan horseradish peroxidase
• Larutan standar feritin.
• Buffer A : Phosphate-buffered saline pH 7,2
• Buffer B : 5 gram BSA (Bovine Serum
Albumin) dalam 1 liter buffer A
• Buffer C : Carbonate buffer pH 9,6
• Buffer D : Citrate phosphate buffer pH 5.
• Larutan substrat
53
54. Feritin Serum
• Melapisi microtitre plate
Antiferitin Ig G + 2
µg/ml buffer C 200 µl 0,05% BSA
dalam buffer C
@200 µl
Tutup dan
inkubasi Kosong
(40C) kan
Cuci tiap sumuran Diamkan (30 menit-
dengan buffer A suhu ruangan)
sampai 3x
54
55. Feritin Serum
20 menit
Microtitre plate reader
50 µl serum 200 µl larutan – 492 nm
pasien + 1 ml standard
30 menit
buffer B
50 µl asam
sulfur 4M
Tutup dan diamkan
(20 menit,-suhu
ruang) Inkubasi 30
menit
200 µl larutan
Kosong kan substrat
Cuci dengan buffer A 3x
200 µl prep.
konjugasi
antiferitin Ig G + Tutup dan diamkan Cuci dengan
HPA diencerkan (2 jam,-suhu ruang) buffer A 3x
55
56. Feritin Serum
Pertimbangan memilih metode assay untuk feritin :
• Deteksi limit : IRMA 10 µg/l, ELISA 1 10 µg/l
• High-dose hook
• Interferen oleh protein non-feritin dalam serum
• Reproducibility
• Dilusi sampel serum
• Akurasi
56
57. IRMA vs ELISA
• IRMA sudah jarang dipakai karena adanya
bahaya radioaktif
• Deteksi limit IRMA lebih besar (10 10 µg/l)
dibandingkan ELISA (1 µg/l)
57
58. Faktor yang mempengaruhi
pemeriksaan status besi
• Diet : rasio enhancer : inducer
• Jenis kelamin : wanita < pria ( haid, partus)
• Umur
• Aktivitas fisik (Ferritin >> ok kerusakan otot dan
inflamasi)
• Siklus menstruasi
• Kehamilan
• Kondisi lingkungan
• Variasi diurnal : lebih tinggi pada pagi hari
dibanding malam hari – berdasar variasi pelepasan
besi dari RES ke plasma
58
59. Prussian-blue staining
• Prussian-blue staining can be applied to films that have
previously been stained by Romanowsky dyes, even after
years of storage.
• It is advisable to let the films stand in methanol overnight to
remove most of the Romanowsky stain. The film should be
checked before carrying out Perls' reaction to ensure that
there is no residual blue staining that could obscure Prussian-
blue staining.
• Sundberg and Bromann described a technique whereby films
were stained first by a Romanowsky dye (Wright's stain) and
then overstained by the acid-ferrocyanide method.
• This can give beautiful pictures, but the small blue-stained
iron-containing granules tend to be masked in young
erythroblasts by the general basophilia of the cell cytoplasm.
Hayhoe and Quaglino described a method for combined
periodic acid–Schiff (PAS) and iron staining.
• This may be helpful in the investigation of abnormal
erythropoiesis in which the erythroblasts give a positive PAS
reaction
59
• Iron-containing granules stain dark purple.
60. Prussian-blue staining (Pearl’s reaction) on aspirated bone
marrow particles to demonstrate iron stores.
A. Normal B. Absent C. Increased D. Grossly increased
60
61. Pathological sideroblasts. There is massive
accumulation of iron-containing granules in
normoblasts and phagocytic cells. Perls' reaction
61
62. Pathological sideroblasts. Sideroblastic anaemias.
Accumulation of iron-containing granules in
normoblasts, arranged characteristically around the
nucleus. A: Hereditary type;
62
63. Pathological sideroblasts. Sideroblastic anaemias.
Accumulation of iron-containing granules in normoblasts,
arranged characteristically around the nucleus B:
myelodysplastic syndrome. Perls' reaction.
63
64. DEMONSTRATION OF HAEMOSIDERIN IN URINE
• Centrifuge 10 ml of urine at 1200 g for 10-15 min.
• Transfer the deposit to a slide, spread out to
occupy an area of 1-2 cm, and allow to dry in
the air.
• Fix by placing the slide in methanol for 10-20 min
and then stain by the method used to stain blood
films for siderocytes (
• Haemosiderin, if present, appears in the form of
isolated or grouped blue-staining
granules, usually from 1-3 µm in size, they may be
both intracellular and extracellular. If
haemosiderin is present in small amounts, and
especially if distributed irregularly on the slide, or if
the findings are difficult to interpret, the test
should be repeated on a fresh sample of urine
collected into an iron-free container and
centrifuged in an iron-free tube. 64
66. Hemosiderin
GRADASI KRITERIA KANDUNGAN BESI
(µg/g)
0 Tak tampak granula besi 43 ± 23
1 Granula-granula kecil pada sel retikulum 130 ± 50±
tampak hanya dengan minyak emersi
2 Sedikit granula-granula kecil yang dapat 223 ± 75
dilihat dengan lensa kekuatan lemah
3 Sejumlah granula-granula kecil dalam 406 ± 131
semua partikel sumsum tulang
4 Granula-granula besar dalam kelompok 762 ± 247
kecil
5 Granula-granula besar dalam kelompok 1618 ± 464
besar
6 Deposit yang sangat besar yang 3681 ± 1400
mengaburkan gambaran sumsum tulang
secara rinci
66
67. %Transf.
Disease SI TIBC Ferritin
Saturation
Iron Deficiency Low High Low Low
Hemochromatosis High Low High High
Normal/
Chronic Illness Low Low Low
High
Normal/
Hemolytic Anemia High High High
Low
Sideroblastic Normal/ Normal/L
High High
Anemia High ow
Iron Poisoning High Normal High Normal
67
68. Hepcidin
• Hepcidin is a small peptide of 25 amino
acids, which is cleaved from a larger precursor.
• It is produced in the liver, but it is present in the
plasma and excreted through the kidneys.
• Hepcidin is structurally similar to antimicrobial
peptides involved in innate immunity, and it has
been shown to have antimicrobial properties.
68
69. Hepcidin
• Hepcidin binds to the iron exporter
ferroportin, causing ferroportin to leave the cell
membrane, enter a lysosomal compartment, and
be degraded.
• Thus, hepcidin directly and coordinately controls
the entry of iron into the serum from absorptive cells
of the intestine and from tissue macrophages.
69
71. Hubungan mol dg massa (gram)
Massa molar adalah massa satu mol zat yang
dinyatakan dalam gram.
Rumus massa adl
Gram = mol x Ar atau Mr
Ar = Massa atom
Mr = Massa molekul / Berat Molekul (BM)
Massa atom (Ar) Fe = 56
Massa molar Fe = 56 gram
(satu mol Fe mpy massa 56 gram)
• Berapa gram massa 10 mol Fe (Mr = 56)?
Massa Fe= 10 mol x 56 = 560 gram
71
72. Interference Serum Iron
• Tembaga dalam serum
• Hb 100 mg/dL 6,9% positive interference in 13,1
µmol/L iron sample
• No significant icteric or lipemic interference
72
73. Iron and TIBC
Interference
• Use of chloramphenicol and hormonal
contraceptives can cause falsely elevated test
results
• Corticotropin can produce falsely low test results
• Iron supplements can cause falsely elevated serum
iron values but falsely low TIBC
• Check the patient history and withhold such drugs
as chloramphenicol, corticotropin, iron
supplement, and hormonal contraceptives, as
ordered. If such medications must be
continued, note this on the laboratory request
73
78. • Soluble Transferrin Receptor (sTfR)
- akhir2 ini sTfR banyak dipakai untuk
memperkirakan cadangan Fe tubuh .
- TfR = mol. Transmembran yg diekspresi
kan pada permukaan semua sel yang
memerlukan Fe . Sebagian TfR dapat
lepas ke sirkulasi → terukur sebagai
sTfR
78
79. - Kadar sTfR menunjukkan kebutuhan
kebutuhan Fe dari jaringan eritropoitik
→ pada peningkatan eritropoisis
maupun Defis-Fe berat → me↑ sTfR .
- Penentuan sTfR : turbidimetrik atau
nephelometrik, tapi belum ter
standardisasi .
79
80. - Berbeda dgn ferritin, selama fase
defisiensi-Fe, kadar sTfR tetap stabil
dan baru bila pe↓ Fe menghambat
eritropoisis, kadar sTfR akan ↑ .
- sTfR = marker serum pendeteksi Fe-
deficient erythropoiesis .
- sTfR tdk berubah pada fase-akut dan
kehamilan → sensitivitasnya lbh baik
daripada saturasi-transferrin
80
81. SERUM TRANSFERRIN RECEPTOR
• The transferrin receptor consists of two
identical, protein subunits of molecular mass 95 kDa.
• The nucleated red cells in the bone marrow, which
synthesize haemoglobin, have the greatest number
of transferrin receptors.
• Transferrin receptor synthesis is also controlled by iron
supply. The mechanism involves IREs , iron regulatory
elements at the 3′ untranslated region of the receptor
mRNA.
• In the presence of adequate iron
concentrations, binding of iron by the IRP changes
the conformation of the protein and prevents its
binding to the mRNA. The mRNA is rapidly broken
down, and synthesis of transferrin receptors is
reduced 81
82. SERUM TRANSFERRIN RECEPTOR
• In 1986, Kohgo et al reported that transferrin
receptors were detectable in the plasma by
immunoassay. Since then, there has been much
investigation of the physiological and diagnostic
significance of circulating transferrin receptors.
• The protein is derived by proteolysis at the cell
membrane and circulates bound to transferrin.
Plasma concentrations reflect the number of
cellular receptors and, in patients with adequate
iron stores, the number of nucleated red cells in the
bone marrow.
• Because the number of cellular transferrin receptors
per cell increases in iron deficiency, concentrations
also increase when erythropoiesis becomes iron
limited.
82
83. SERUM TRANSFERRIN RECEPTOR
• There has been no agreement about the source of
transferrin receptor as standard or as an antigen for
the raising of antibodies. Transferrin receptors have
been purified from placenta and from serum.
• No “reference” method is therefore described here.
Three enzyme immunoassay kits (Orion; Ramco; RΔ)
for the determination of serum transferrin receptor
concentrations have been evaluated for the Medical
Devices Agency. All have been approved for
diagnostic purposes in the United States by the Food
and Drug Administration. Assays for fully
automated, diagnostic, immunoassay systems are
now being introduced and offer improved
sensitivity, reproducibility, and speed.
83
84. SERUM TRANSFERRIN RECEPTOR
• The different reference ranges in the available
commercial assays reflect the differences in preparations
of transferrin receptor used to raise antibodies and as a
standard in the various assays.
• For the Orion, Ramco, and RΔ kits Akesson et al and
Worwood et al noted some assay drift but found
acceptable intraassay coefficient of variance (CV)
values.
• The determined sensitivity was adequate for clinical
purposes for the three assay systems. There are
differences in units and absolute amounts for serum
transferrin receptor concentrations.
• Four different units (nmol/l, g/ml, mg/l [ng/ml], and kU/l)
and different normal ranges are in use.[59] At the present
time serum transferrin receptor (sTfR) is not included in the
national external quality control schemes for the UK
(NEQAS) and the Welsh External Quality Assessment
Scheme (WEQAS).
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