CORD

BLOOD
BANKING
Dr Vasanthakumar Gounder

Moderated by

Prof Dr Ratti Ram Sharma

 • Cord blood is the remaining blood in the placenta
and the umbilical cord after the child-birth.
• It is being collected and stored to treat various
disorders.

What is
• Topics to be discussed
• Cord blood – History
• Licensing of Cord Blood Bank
• Contents of the Cord Blood

Cord
• Collection process and storage
• Diseases can be treated
• Types of cord blood transfusion

Blood?
• Public vs Private cord bank
• Advantages of cord blood over PSC
• Recent advances in Cord Blood Bank
• Research papers from our institute
 • Dr. Eliane Gluckman – 1988
• Dr. Auerback – clinician.
• The first cord blood transplantation in Fanconi

History
Anaemia patient – Mathew.
• First UCB Bank in new York – Professor Harold
Broxmeyer in 1991
• First unrelated UCB bank was established by Dr.
Pablo Rubindtein in 1992

Reference:
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC544272
3/
Cord Blood Bank.
• Every year, 10,000 children are born with thalassemia and every year over a lakh people are
identified with leukaemia.
• If they were to look for HLA match for treatment, there is no inventory of any great size in India
yet.
• HLA match is ethnicity dependant. When an Indian is looking for a match there is a greater
likelihood of finding a match within an Indian inventory.
• Jeevan Blood Bank statement - Our goal is to have 30,000 donated cord blood units and 1,00,000
bone marrow donors registered by 2022. This will enable 70 percent of Indians with blood cancers
to find a match and hope for a cure.
• Reference: https://www.thehindu.com/news/cities/chennai/After-21-years-Jeevan-Blood-Bank-
shuts-shop/article17151728.ece
Stats about cord blood banking.
• With India's booming birth rate of 26 million births per year and genetic diversity; the country would be
poised to be the largest collector of UCB in the world.
• Four banks are established in India – Relicord, Jeevan Cord, and Stemcyte, School of Tropical Medicine,
Calcutta.
• Collectively having 5,000 units. Similarly, seven private banks have been established to date. These are Life
Cell with maximum inventory of 19,000 followed by Cryo Banks having 17,000 plus samples and about
4,500 between Cryosave, Cord Life, Baby Cell, Stem One, and ISSL (International Stem Cell Service)
(personal communication from Dr. Phagun Shah, Medical Advisor, Cryobank, India).
• The highest inventory of 48,808 UCB is with New York Blood Centre's National Cord Blood Program. This
type of program in general is funded in part or entirely by public funds
CORD BLOOD
BANK LICENSING
Licensing.
• Umbilical Cord Blood banks (UCB) are permitted only
under license and monitoring by the Central Drug
Standards Controlling Organization (CDSCO).
• The Cord Blood Banks have to comply with the Drugs
and Cosmetics (3rd Amendment) Rules, Gazette
Notification No. GSR 899(E) dated 27/12/2011 for
collection, processing, testing, storage, banking, and
release of stored units.
• 28-F for processing of cord blood bank.
• (Available at: http://cdsco.nic.in/html/GSR%20899.pdf).

• GUIDELINES -
http://www.arthiqs.eu/assets/user/files/CBB/D9%20CBB
%20Guide%20%20jan%202018.pdf
MOMENT PLEASE!!

What is the form number that is used

for permission of processing WB and
preparation of blood components?
Facts to be remembered!!

• 27 – C Granting/renewal for the processing of WB and preparation of Blood

Components.
• 28- C Blood bank products for sale/distribution.
• 26- G Renewal of Blood Bank License.

• 28- F Permission for processing cord blood bank.

• 26 – J Cord blood renewal.

• Reference: https://cdsco.gov.in/opencms/opencms/en/FAQ/
NICE TRY!!
Many more Qs to
attempt.
CRITERIA
Are Hematopoietic Stem Cells Nucleated?

Haan Ji.

They are round, non-

adherent, with a rounded
nucleus and low
cytoplasm-to-nucleus ratio.
• Cord blood is collected from full-term deliveries and can be collected
either in utero or ex utero.
• In utero cord blood collections are obtained by a trained medical
professional during the third stage of labour. A standard 250 mL blood bag
with CPD anticoagulant is used and cord blood drains into the bag under
gravity.
• Ex utero cord blood collections are obtained after the delivery of the
placenta which is suspended above the collection bench and once again
blood drains into the collection bag under gravity.
• Both methods of cord blood collection produce good quality cord blood
units as long as the cord blood is collected by properly trained staff.
• The total nucleated cell count (TNC) of a cord blood unit has been
found to be critical in the overall success of subsequent transplantation.
(Cord blood contains RBCs, Platelets, and Plasma – we’re not interested in
these non-nucleated structures. They are later discarded. We are looking for
Stem Cells, which are nucleated and found in the buffy coat layer. Stem
Cells comprise only 1-2% of Total Nucleated Cells).
• Cord blood was traditionally collected and frozen whole (without any
processing) but it soon became clear that such a practice would in the long
term require far too much storage space.
• Most cord blood banks now reduce the volume of the cord blood unit to a
fixed volume of buffy coat layer which contains the stem cells needed for
transplant.
• The red cells and plasma are discarded. (i.e. we only keep buffy coat)
• It is very important that these volume reduction processes retain maximum
numbers of TNC and CD34 + myeloid progenitor cells.
• The volume reduction technology was originally a manual process
involving the ‘open’ manipulation of the cells using hydroxyethyl starch
(HES). Automation was first introduced in 1999.
• Volume reduction also allows for the use of lower volumes of the
cryoprotectant DMSO which allows the transplantation of volume-
reduced cord blood units without any washing procedures.
• Standard cryopreservation protocols using DMSO at a concentration of 10% and a
controlled rate freezer followed by storage in liquid nitrogen at or below −135 °C result in
an average of 80% recovery of TNC and > 90% recovery of CD34 + myeloid progenitor
cells.
• All cord blood donated or collected for public transplantation must be screened for
infectious disease, typed for HLA and ABO Rh typed.
• All cord blood banks operate under either local regulation (e.g. the Human Tissue
Authority in the UK) or international regulation (e.g. the American Association of Blood
Banks, Association for the advancements of blood and biotherapies AABB) to ensure that
cord blood unit for transplantation are produced to the appropriate standard.
• In the early days of cord blood stem cell technology, cord blood was collected for
unrelated allogeneic use only.
• It then became clear that there was a role for ‘directed’ donation of cord blood where there
is a disease in the family that may be treated using cord blood stem cells. Such cord blood
collections are carried out at the request of the physician treating the patient and with the
agreement of the obstetric team caring for the mother.
• Related cord blood stem cell transplantation is now the first-line treatment in patients
suffering from thalassemia major. The use of prenatal genetic diagnosis (PGD) also allows
the collection of cord blood from HLA identical siblings.
COMPATIBLITY – what all is checked
•HLA Compatibility
•Total Nucleated count
•CD34+ count- Marker protein
•Stem cells – Account for only 1-2% of TNC.
•Colony forming unit – the ability to function and multiply,
growth media, and the colonies are counted.
•MNC- Mononuclear cells – Single nucleus/ can be identified
how it multiplies
HLA Matching
1. HLA‐matching of the recipient and
CB Unit.
2. Collected cell dose (TNC ± CD34+)
3. Patient’s diagnosis
(malignant versus non‐malignant)
4. Avoiding CBU containing antigens
that match with the specificity of any pre‐
transplant donor‐specific anti‐HLA
antibodies in the recipient. (obviously,
like crossmatch)
1) HLA‐matching
HLA‐matching should be based upon allelic typing for HLA‐A, ‐B, ‐C and ‐DRB1 for single UCBT
1.Select an HLA‐matched (8/8) CBU. TNC dose should be >3 × 107/kg
2.If an HLA‐matched (8/8) CBU is unavailable; select a CBU matched at 7/8 HLA ‐loci. HLA‐A or HLA‐B mismatches are preferable
to HLA‐DRB1 mismatches. TNC dose should be >5 × 107/kg for 5–7/8 matched units
3.If a CBU matched at 8/8 or 7/8 HLA‐loci is unavailable, consider a CBU matched at 5 or 6/8 HLA ‐loci. Avoid mismatches in HLA ‐
DRB1.
4.4/8 matched CBU may rarely be considered as a single CB graft if no other option is available. TNC dose should be >5 × 107/kg for
4/8 matched units
5.CBU 3/8 HLA matched CBU is not recommended

(2) TNC and CD34+ cell dose Non‐malignant disorders

Malignant disorders
Total Nucleated cell dose
Nucleated cell dose At freezing, minimum cell
At freezing, the minimum TNC dose 3·5 × 107/kg, or
dose 3·0 × 107/kg, or After thawing, minimum
After thawing, minimum TNC
of 2·0–2·5 × 107/kg*
cell dose 3·0 × 107/kg

CD34+ cell dose**
At freezing, 1·0–1·7 × 105/kg, or
After thawing, around 1·0–
1·2 × 105/kg
For patients with BM failure syndromes (aplastic anaemia or
congenital bone marrow failure states) or
haemoglobinopathies, the number of TNC at freezing should
be greater than 5 × 107/kg
CD34+ cell dose At freezing or after thawing,
>1·7 × 105/kg
3) Other considerations when selecting single CB units
If many CBU meeting the criteria above are available, the following factors should also be considered:
•Use accredited Cord Blood Banks. For safety, only accredited banks recognized by national and international organizations
should be used
•ABO compatibility: ABO compatibility may be associated with improved outcomes, although the data are conflicting
•Sex matching: Sex matching between CBUs and patients in single or double UCBT is not necessary
 3ml samples are taken for

TESTING • HLA typing,

• Nucleated cell (NC) count,

OF THE
• CD34+ cell count,
• Transmissible diseases screening (TTI
Screening)

CORD • Genetic testing.

• Tests for fungal, aerobic and anaerobic
bacteriology cultures.

BLOOD • ABO AND Rh matching

CORD BLOOD
COLLECTION
PROCESS
Collection process
• Clean the cord site 4-6 inches above and below with the disinfectant
• The needle is inserted to draw the blood
• The bag has to be kept at a lower level to facilitate the gravitational flow
• Bag has to be mixed at a regular interval to promote the mixing of anti-coagulant (CPD) with the cord
blood.
• Around 80-120 ml of blood is collected.
• The process is performed in a closed system with the use of sterile connecting devices to reduce bacterial
infections due to contamination.
• The NK (Natural Killer) and CD34+ cell estimations are repeated using the pelleted fraction so that there is
a monitoring of the quality of the process.
• WBCs present in the collecting bag are transferred to a freezing bag – for calculation of CD 34 count on
them.
At the end of the freezing procedure the cells are stored in liquid nitrogen freezer.

Immunophenotyping of the CD34+ cell estimation is carried out on the whole blood before starting the
processing and the volume reduction.

The CD34+ cell number is calculated on the WBC units present after the processing.

In vivo - as soon as the baby is delivered, CB is collected.

Ex-vivo- the placenta is being taken out with the clamped cord and the cord blood is collected.
 MOVIE
TIME

CORD
BLOOD
PROCESS
Cord blood
collection- • Link - https://youtu.be/Mwm6gvubLrY

Macopharma
video.
 • The UCB unit will be transferred to a 150-ml bag and
the HES solution will be directly added to the
collection bag under sterile conditions.
• Centrifugation – to remove the plasma
• Sedimenting agents - to remove the red blood cells
Hydroxyethyl starch, Gelatin,
Preparation polygeline, and dextran
• Cryoprotective solution – 10% Dimethyl Sulfoxide

and processing (DMSO)

• Slow cooling @ controlled rate freezing 1deg C/ Min
• AABB recommends the temp should be < -150deg C
• Once frozen, the bag is usually transferred to a long-
term storage container and immersed in the liquid
nitrogen -196deg C.
 Before processing

QC TO
• TNC
• Hematocrit
• ABO/Rh compatibility

BE
• CD 34+ cell enumeration
• CFU assay
• Viability

DONE: • Sterility
• HLA typing.
CONTENTS OF
THE CORD
BLOOD
• Hematopoietic stem cells (HSC)- myeloid
and lymphoid series.
• Non-Haematopoietic stem cells which are
Endothelial progenitor cells known as
Endothelial Cord Forming Cells (ECFC)
• Present in UCB – 2-5 Cells/ml.
• Used in Ischemia and defective wound
healing to improve neoangiogenesis.
• Stromal Cells – Mesenchymal cells (MSC)
Other than Cord Blood.
• NK cells – These cells kill different targets such as cancer or virally-infected cells without any prior activation.
• Cord bloods’ pluripotent and totipotent stem cells produce organ-specific cells.
• Wharton’s jelly - to heal the wounds as they have the properties like high plasticity, proliferative, differentiation capability,
and also low immunogenicity.
• Wharton’s jelly + poly vinyl Alcohol – to treat chronic skin wounds.
W
FUNCTION
OF HSC
AND MSC
FUNCTIONS
• HSC
• Differentiates into lymphoid and myeloid series
which produces T-cell, B-cell and NK cell and the
latter one forms the RBC, WBC and platelets.
• MSC
• Differentiates into muscle, bone, cartilage, tendon,
fat, and bone marrow stroma
• It prevents the GVHD.
• MSC – a potential therapeutic remedy that controls
the immune cells and prevents inflammation,
thereby reducing the GVHD.
JUST A
MOMENT!!
1.GVHD vs GVL, Which is beneficial to
the patient?
ANSWER

GVL – GRAFT VS
LEUKEMIA REACTION IS
BENEFICIAL TO THE
PATIENT.
 • T cells plays a major role in both ways, they
place a major role in GVL effect, as well in
GVHD.

Importance • When transplanting T-cell depleted stem cell

transplant, GvHD can be partially prevented,
but in the same time the GvL effect is also
of T cells reduced, because T-cells play an important
role in both of those effects.
• There are no methods available to promote
only the GVL and not the GVHD.
TYPES OF
CORD BLOOD
TRANSFUSION
 Types

Double • If single cord blood is not sufficient, we can add

the matched blood, pooled and then transfuse to
umbilical cord the patient.
transfusion

• To have the adequate effects, patient can be

Tandem transferred with one more unit as a successive
transfusion attempt.
 DISEASES
THAT CAN BE
CURED
 • AML – Most common indication for allogenic tx.
• MM – Most common indication for autologous tx.
• ALL- Most recently, a 90% complete remission rate
was achieved in high-risk patients with relapsed or
refractory ALL who received genetically modified
autologous T cells expressing a chi­meric antigen
receptor (CAR-T cells) enabling them to recognize and
kill all CD19-expressing cells, including CD19-

Medical Rx
expressing ALL
• HSCT – is done only in High-risk ALL/Philadelphia
positive cases/MLL mutation

vs Cord • CLL- Bruton’s tyrosine kinase inhibitor, ibrutinib, and

the phosphatidylinositol-3-kinase delta (PI3 Kδ)
inhibitor, Idelalisib.Cellular therapy targeting CD-19 is
Blood Tx also a promising approach
• The question if transplant is still efficacious than a
combination of proteasome inhibitor (e.g., bortezomib)
and an immunomodulator (e.g., lenalidomide) still
remains unans­wered and is the focus of an ongoing
clinical trial (NCT01208662, Clinicaltrials.gov .
 • Around 13 cases of thalassemia have been
treated using UCB transplantation.
• Of these, six cases were transplanted using fully
matched sibling UCB at Apollo hospital,
Thalassemia and Chennai. UCB units were obtained from Life
Cell and Cord bank.

Cord Blood Bank. • Thalassaemia free survival rate was 83%.

• Reference:
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC
3193706/
QUESTION
TIME
1. Can we cure ‘GENETIC’ disorders using autologous
UCBT?
 ANSWER

NO, COZ AUTOLOGOUS

TRANSFUSION WILL CARRY
THE SAME DEFECTIVE GENE.
GOOD!
 OUR
INSTITUTE
RESEARCH
PAPERS
Conclusion:
• https://symbiosisonlinepublishing.com/stem-cell-
research-therapeutics/stem-cell-research-
therapeutics02.php
• A maternal age over 25 years, fetal birth weight > 3 kg, in-
utero collection method, cord clamp- collection interval 1
minute showed better initial cellular yields.
• DMSO concentration of 5 % was found optimum for
cryopreservation for best TNC or MNC recovery and 10
% for best recovery of CD34+ cells.
• Post thaw no wash technique appeared to be better method
as cell loss due to washing was significant.
PUBLIC CORD
BANK

VS

PRIVATE
CORD BANK
 Free

Public Available to
Blood everyone who
needs them
Bank Genetic and
infectious part will
be tested
PRIVATE BLOOD BANK

Costly

Very minimal Only focused on

possible options autologous tx

Can only be used

Not available for
by the family
public
members
 CORD
BLOOD
BANK
• USA has twenty-eight Public cord blood banks and twenty-nine private cord blood banks.
• UK also implemented both Private and Public Cord Blood banking and there are two public Cord blood
banks and six Private cord blood banks.
• South Africa being prominent country in African continent, has three private UCB Banks.
• Singapore a well-developed country in Asian continent has only one public cord blood banks.
• India, as a developing country plays a major role in cord blood banking. India has four public cord blood
banks and fourteen private cord blood banks (Seah, 2014).
Private sector- cord blood banks in INDIA
• Jeevan Stem Cell Bank (1995)
Jeevan (also known as BeTheCure.in) is the first purely public cord blood bank set up in India, funded by a tie-up with the state of Tamil
Nadu, that enables quick and affordable (free for the poor) access to matching stem cells from donated umbilical cord blood for the life-
saving treatment of Indians with blood cancers, thalassaemia, and other blood disorders.
• Reliance Dhirubhai Ambani Life Sciences Center, Thane-Belapur Road, Navi Mumbai
Reliance Life Sciences Pvt. Ltd. established the first cord blood repository in South East Asia and has offers both public donation and
family storage.They are a part of the Reliance Group of companies, a member of the Fortune Global 500.
• The School of Tropical Medicine (STM)
The School of Tropical Medicine (STM) in Kolkata has launched the first public cord blood bank in India that is funded by the government.
The cord blood bank is operated by the school’s Dept. of Regenerative Medicine and Translational Science. The bank was announced in
early 2014 and launched at the end of 2015.
• StemCyte India
StemCyte India is a joint venture between StemCyte Inc., Apollo Hospital Enterprises Ltd., and Cadila Pharmaceuticals Ltd. was established
in 2008, dedicated to collection, processing, testing and storage of both – private and public umbilical cord blood units and its therapeutic
applications.
• Lifecell
Established in 2004, LifeCell is India’s first private stem cell bank, on the occasion of successful preservation of 1,00,000 umbilical cord
stem cell units at its facility in Chennai, announced launch of its umbilical cord public stem cell bank in July 2014, aimed to help patients in
need of a lifesaving stem cell transplant improve their odds of finding matching stem cells.
Recommendations for Cord Blood Unit (CBU) Selection in 2014. Three Criteria are Major for the Choice of CBU: Human Leukocyte Antigen (HLA) Matching, Total
Nucleated Cell Dose at Freezing, and Patient HLA Antibodies

Human Leukocyte Antigen (HLA) Match

l Compatibility 6/6, 5/6, or 4/6
l Antigenic HLA-A and B, allelic DRB1
l Prefer HLA-A and B mismatches to DRB1 l For nonmalignant diseases:
l Matching is particularly critical and DRB1 mismatch should be avoided
l Double cord blood transplantation (CBT): l 4/6 minimum between the two units
Total Nucleated Cell (TNC) Dose at Freezing
l Malignant diseases: ≥3 × 10e7 TNC/kg
Compatibility 6/6 or 5/6
l Nonmalignant diseases: ≥3.5 × 10e7 TNC/kg
l Malignant diseases: ≥3.5 × 10e7 TNC/kg
Compatibility 4/6
l Nonmalignant diseases: ≥4 × 10e7 TNC/kg
⇛ If these cell doses are not met with one unit, a double CBT should be performed
≥2 × 10e7 TNC/kg for each unit
Anti-HLA Antibodies
Exploration of anti-HLA immunization by a sensitive method for all recipients
⇛ Units against which there are a very intense antibody response should be particularly avoided
Other Criteria for CB Selection

When faced with multiple cord blood units (CBU) that

fit the above criteria, the following criteria should be considered:
l HLA high resolution typing: Matching for HLA-C (-DQB1)
and for HLA class I alleles should be included in the CBU selection HLA mismatch in the “GvH direction only” should be preferred.
l Dose of CD34+ cells ≥1.5 × 105/kg
l ABO matching: The use of ABO compatible units should
be taken in consideration
l Noninherited maternal antigen match: Select an NIMA-
matched CBU?
ADVANTAGES OF CORD BLOOD
OVER PERIPHERAL STEM CELL
TRANSPLANTATION
•Easy procurement.

•No risk or discomfort to donors.

•Low incidence of viral contamination (38.2%) compared to BM

•Able to be stored at cryogenic temperatures indefinitely without significantly affecting cell viability.

•Immediately available and easily shipped.

•There is a high immune tolerance of UCB cells because they are unable to generate cytotoxic T-Lymphocytes, which
respond to allogenic antigens. UCB cells are also unable to synthesize the proinflammatory cytokines interferon- γ (IFN- γ)
and tumor necrosis factor-α (TNF-α)

• UCB has an abundant source of both hematopoietic and nonhematopoietic stem/progenitor cells, which is comparable to
BM.

•There is a large number of colony-forming unit-granulocyte macrophage (CFU-GM) compared to adult sources

•Procurement and use are not associated with the same ethical considerations as embryonic and fetal tissues.
Disadvantages
• Future usage: There is a 1 in 217 chance that your child will need a stem cell transplant. There are chances that you may
never need those preserved stem cells. However, having a useful resource saved in case of any misadventure avoids
unnecessary delays and expenses.
• Limitation in the treatment of diseases: There exist limitations in terms of what all medical conditions can be treated by
stem cells. For instance, if your child is born with a genetic condition then the stem cells will also carry that faulty gene.
Therefore, a child cannot use his or her own stem cells in that case. This is a major drawback of private banking wherein
you are only allowed to use your own child’s stem cells. However, with community stem cell banking, you can access a
large pool to find a donor match.
• Storage duration: Cord blood banking came into existence around 25 years ago. Therefore, there is no complete
information on how long these stem cells can be preserved in a viable condition. However, researchers believe that if
the best preservation practices are followed, stem cells can be effectively stored for many decades.
• Disposal: Private stem cell banks can discard preserved cord blood stem cells when payment isn’t made or the family no
longer wants to preserve the stem cells.
• Cost: Parents are often in a fix because of the high costs charged by private stem cell banks. However, LifeCell
community banking offers affordable storage plans and reasonable EMIs for parents, starting at just INR 950/month…
(LOLs)
• Additional blood requirement: Delay in cutting the child’s umbilical cord can allow blood to flow back to the child which
can reduce the risk of iron deficiency. Parents can opt to delay clamping by 30 to 60 seconds as recommended by the
American College of Obstetricians and Gynaecologists (ACOG).
 • The major problem faced in India is the
collection of UCB due to high cost and

What’s the
comparatively less functional public banks. In
addition, considering a large population with
deliveries in public hospitals due to low cost,
UCB storage in India needs an increased
final public-private partnership model where UCB
can be stored by affordable and non-
affordable people as well.

outcome? Is • Even though it is beneficial in many

conditions, still people need awareness in this

it worth?
area.
• Since, this is being considered as an
alternative option, it will take some years to
be implemented completely.
TAKE HOME
MESSAGE
LONG WAY TO
GO!!!

Licensing

Genetic diseases can’t be treated.

HLA DRB1 matching matters than HLA A & HLA B

TNC count should be more than 3*107

CD4+ count should be in the range of 1.5-2.0

It’s been 25 years since the first cord blood transfusion,

but still, a long way to go!
Happy Birthday Dr.
Lakhvinder Sir.