dr r v s n sarma md msc consultant physician & chest specialist
DESCRIPTION
Stem Cell Research Advancements & Applications Special focus on Cord Blood Basics of Cloning and GE. Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist. www.drsarma.in. The Outline of Discussion. What are Stem Cells ? Why Stem Cells ? What are the sources of Stem Cells (SC) ? - PowerPoint PPT PresentationTRANSCRIPT
![Page 1: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/1.jpg)
www.drsarma.in1
![Page 2: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/2.jpg)
www.drsarma.in22
Dr R V S N Sarma MD MSc
Consultant Physician &
Chest Specialist
www.drsarma.in
Stem Cell Research
Advancements & Applications
Special focus on Cord Blood
Basics of Cloning and GE
Stem Cell Research
Advancements & Applications
Special focus on Cord Blood
Basics of Cloning and GE
![Page 3: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/3.jpg)
www.drsarma.in3
The Outline of Discussion
What are Stem Cells ? Why Stem Cells ? What are the sources of Stem Cells
(SC) ? What are the methods of SC
Production ? How Cord Blood SCs are harvested ? What are the applications of Stem
Cells ? Where are we now ? Where do we go from here ? Cord Blood Banking resources –
![Page 4: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/4.jpg)
www.drsarma.in4
Cell Therapy – Regenerative Medicine
Autologous (from the same individual) Allogeneic (from a different
individual) Xenogeneic (from a different species)
1. Differentiated – specialized cells of same or other tissue type, such as heart, muscle, blood cells etc.
2. Undifferentiated or unspecialized or uncommitted cells such as stem cells.
![Page 5: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/5.jpg)
www.drsarma.in5
Different Types of Stem Cells
Totipotent (totally undifferentiated – potential to become any cell type including whole organism)
Example: The inner cell mass of embryo
Pluripotent (undifferentiated – potential to be any cell type but not an entire organism)
Emb Stem Cells (hESC), Emb Germ Cells (hEGC)
Multipotent (differentiated and yet undifferentiated)
The UCSC, BMSC, MSC – limited in potential
Use of the stem cells raises ethical / legal concerns
Wide variation in national policies for stem cell use
![Page 6: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/6.jpg)
www.drsarma.in6
Hierarchy of Stem Cells
Multipotent
![Page 7: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/7.jpg)
www.drsarma.in7
What is special about Stem Cells ?
1. Capability of dividing and renewing themselves for long periods - almost immortal
2. Stem cells are unspecialized “uncommitted” cells.
3. Stem cells can give rise to specialized cells. Remain “uncommitted” until they receive signals from their environment to develop into specialized cells.
4. Stem Cells have Plasticity - Stem Cells from one tissue are able to give rise to cells of a completely different type of tissue
![Page 8: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/8.jpg)
www.drsarma.in8
Hectic Competition
The ultimate in love
![Page 9: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/9.jpg)
www.drsarma.in9
Week 1 of Gestation
FertilizationDay 1
2 cell stage Day 2
4 cell stage Late Day 2
MorulaDay 3
BlastocystDay 5
ImplantationDay 6 - 7
![Page 10: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/10.jpg)
www.drsarma.in10
The Blastocyst – Day 5
The Blastocyst consists of
Inner Cell Mass
(embryoblast)
Trophoblast
Blastocele
![Page 11: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/11.jpg)
www.drsarma.in11
Week 2 of Gestation - Day 7.5
Blastocystic Cavity
Cytotrophoblast
Inner Cell Mass
Syncytiotrophoblast
Uterine Epithelium
Uterine Artery
Uterine Gland
Syncitio-trophoblsat
![Page 12: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/12.jpg)
www.drsarma.in12
Week 2: Early Development
Cytotrophoblast
Inner Cell Mass :
Epiblast
Hypoblast
Amniotic Cavity
Amnioblasts
Primary Yolk Sac
Exocoelomic Membrane
![Page 13: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/13.jpg)
www.drsarma.in13
Embryonic Cell Types – at Gastrulation
![Page 14: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/14.jpg)
www.drsarma.in14
GENES
![Page 15: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/15.jpg)
www.drsarma.in15
Sexual Reproduction
![Page 16: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/16.jpg)
www.drsarma.in16
Cloning or Asexual Reproduction
SCNT
![Page 17: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/17.jpg)
www.drsarma.in17
Parthenogenesis
![Page 18: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/18.jpg)
www.drsarma.in18
Embryonic Stem Cell Cultures
![Page 19: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/19.jpg)
www.drsarma.in19
Human Genetic Engineering
![Page 20: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/20.jpg)
www.drsarma.in20
Germline Engineering
Combines the application of
1. Stem Cell use
2. Genetic Engineering
3. Embryo Cloning and
To Produce Designer Babies
![Page 21: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/21.jpg)
www.drsarma.in21
In Vivo – Pluripotent Stem Cells
![Page 22: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/22.jpg)
www.drsarma.in22
Different Types of Stem Cells
Embryonic cells – by In Vitro Fertilization (IVF)
ESC and Embryonic Germ Cells (EGC)
Somatic cell nuclear transfer (SCNT) - Cloning
Used to clone ‘Dolly’
Adult Stem Cells (ASC) – Best eg. BMT, MSC
Umbilical cord blood stem cells (UCSC)
Already differentiated to some degree ?
Can they be made to become pluripotent ?
Will they rapidly lose capacity to differentiate ?
Amniotic Fluid Stem Cells – AFSC – 9th Jan 2007
![Page 23: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/23.jpg)
www.drsarma.in23
ESC versus ASC
Embryonic Stem Cell (ESC)
High malleability Potential for undesired
development (teratomas)
Infinite lifespan,
unlimited supply High ethical burden Uncertain legal status
Adult Stem Cells (ASC) Limited developmental
potential Better behaved, easier
to manage Lose their ability to
proliferate or differentiate after a time in culture
Less moral ambiguity Less legal controversy
![Page 24: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/24.jpg)
www.drsarma.in24
Methods of Stem Cell Generation
SCNT
IVF
![Page 25: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/25.jpg)
www.drsarma.in25
Stem Cell Maturation in Bone Marrow
![Page 26: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/26.jpg)
www.drsarma.in26
Alternatives to Embryonic Stem Cells from Living Human Embryos
From dead embryos Less than 8 wks MTPs Non-destructive biopsy Bioengineered cells (genetically
altered) Reprogrammed adult somatic cells
[but] Live human embryos hold most promise
![Page 27: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/27.jpg)
www.drsarma.in27
Stem Cell Programming Stem Cell Differentiation
‘RECIPE’
![Page 28: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/28.jpg)
www.drsarma.in28
Therapeutic Applications of Stem Cells
Regenerative medicine – permanent repair of failing organs - e.g.Cardiomyocytes for heart diseaseAngiogenesis in CAD – ‘Auto bypass’ Islet cells for diabetesNeural cells for Parkinson’sBlood cells for cancerChondrocytes for osteoporosisKeratinocytes for burns
![Page 29: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/29.jpg)
www.drsarma.in29
Stem Cell Applications
![Page 30: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/30.jpg)
www.drsarma.in30
The Promise of Stem Cell Research
![Page 31: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/31.jpg)
www.drsarma.in31
Innovations of Cell Therapies
![Page 32: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/32.jpg)
www.drsarma.in32
Cell Therapies – Global Over View
![Page 33: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/33.jpg)
www.drsarma.in33
Estimated Market for CT versus IT
![Page 34: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/34.jpg)
www.drsarma.in34
Electron Micrographs of Stem Cells
![Page 35: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/35.jpg)
www.drsarma.in35
Programming of
Stem Cells – ex vivo
EMG of Stem Cell
![Page 36: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/36.jpg)
www.drsarma.in36
Stem Cells in CAD
Cardiomyogenesis Coronary Angiogenesis
![Page 37: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/37.jpg)
www.drsarma.in3737
The Enormous Power of the Umbilical Cord
Let us Understand Let us Understand
![Page 38: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/38.jpg)
www.drsarma.in38
Umbilical Cord Blood Banking
Collection, Processing, Cryo-preservation of blood in the placenta and umbilical cord after the separation of baby
In-Utero collection, Ex-Utero collection
45 disease are now treatable with Cord blood Stem Cells
These are not ‘experimental’ but ‘mainstream’ therapies
The stem cell concentration is 10 times higher than BM
For BMT overall only 25% get a suitable HLA match
Reduced GVH-Disease in the recipient
Painless and easy to collect, Immediately available
BMT costs 3-4 lakhs – Cord blood has lower cost
![Page 39: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/39.jpg)
www.drsarma.in39
UCSC Rx. for which diseases ?
Acute and Chronic Leukemias Myelodystrophic syndromes Stem Cell disorders – FA, PNHU, Aplastic Anemia Myelo and Lympho proloferative Disorders Liposomal storage Diseases, Plasma cell
disorders Phagocyte Disorders Histiocytic Disorders, Platelet abnormalities Other Malignancies – BC, Ewings, NB, RCC RBC Abnormalities – Sickle cell, Beta
Thalassemia SCID – Congenital immune system disorders
![Page 40: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/40.jpg)
www.drsarma.in40
Promising Applications of UCSC
Parkinsonism Alzheimer's Disease Type 1 Diabetes Mellitus Cardio-myopathies Coronary revascularization for CAD Retinoblastoma – Retinopathy Burns and skin regeneration Osteo and other degenerative
arthritides Cirrhosis, Hepatitis Osteoporosis
![Page 41: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/41.jpg)
www.drsarma.in41
![Page 42: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/42.jpg)
www.drsarma.in42
![Page 43: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/43.jpg)
www.drsarma.in43
The Placental Barrier
![Page 44: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/44.jpg)
www.drsarma.in44
Why Use Cord Blood Stem Cells ?
Absolutely non-controversial Absolutely simple to collect More potent than Adult stem cells Non-contaminated Readily available Both for vaginal and c-section
deliveries Low rates of rejection for transplants Most importantly very high chances of
HLA matching
![Page 45: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/45.jpg)
www.drsarma.in45
Stem Cell therapy centres in India
Tata Memorial Hospital, Mumbai Adyar Cancer Centre, Madras Apollo Specialty Hospital, Madras, Apollo Hospital, Global Hospitals, NIMS,
Hyderabad Christian Medical College, Vellore Narayana Hruduyalaya , Bangalore R&R Army Hospital, New Delhi AIIMS , New Delhi Inlaks Hospital, Pune Armed Forces Medical College, Pune Sanjay Gandhi PGIMS, Lucknow
![Page 46: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/46.jpg)
www.drsarma.in46
Storage of Umbilical Bloodfor Stem Cells
Commercial enterprise (Cordbank) stores umbilical stem cells from birth for future autologous use.
“Saving your baby’s umbilical cord stem cells could save your baby’s life”
Others advocate for a public cord blood bank
![Page 47: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/47.jpg)
www.drsarma.in47
Cord Blood Collection
![Page 48: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/48.jpg)
www.drsarma.in48
Thawing Method
Based upon COBLT (Cord Blood Transplantation Study) Method
Used for thawing cord blood products received from COBLT Banks frozen in MedSep (80/20) bags
![Page 49: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/49.jpg)
www.drsarma.in49
Cord Blood Preservation
Under liquid N2 at – 192o C
![Page 50: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/50.jpg)
www.drsarma.in50
Preparation
2 Hours prior to thaw: Review product paperwork
Gather supplies and
reagents
Prepare wash media
(Dextran 40 and human
serum albumin).
Prepare wash bag setup:
MedSep Transplant Set+
300 mL plastic transfer
bag.
![Page 51: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/51.jpg)
www.drsarma.in51
Preparation
Bag 1 Transfer 250 mL Dextran 40 into 300 mL transfer bag. Add 50 mL 25% Albumin. Final conc. of albumin is 4.1%
Connect the Dextran/Albumin bag to a MedSep Transplant Set (Bag 2 & 3) using sterile connecting device. Bag 1 Bag 3Bag 2
![Page 52: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/52.jpg)
www.drsarma.in52
Preparation of Wash Bags
Transfer 125 mL
of the
Dextran/Albumin
to Bag 2 and chill
bags for
approximately
one hour in
refrigerator.
![Page 53: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/53.jpg)
www.drsarma.in53
Retrieval
Remove from
storage
Verify all label
information
with 2nd
technologist
Detach segment
and store
![Page 54: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/54.jpg)
www.drsarma.in54
Thawing
Place the
product inside
a Ziploc bag,
seal tightly,
and submerge
in the 37C
waterbath until
slushy.
![Page 55: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/55.jpg)
www.drsarma.in55
Connect to Wash Bags
Clean scissors and CBU bag
with alcohol. Cut port and
rewipe with alcohol
Connect CBU
With all clamps closed
spike both ports of the
CBU bag with MedSep
transplant set. Open
clamps for CBU and Bag
2 (125mL Dex/Alb
solution)
Bag 2Bag 2
![Page 56: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/56.jpg)
www.drsarma.in56
Diluting Cells
Dilute Cells
Gradually (over 4-
5 minutes) mix the
cells in the
cryobag with the
Dextran/Albumin
solution (Bag 2)
by raising and
lowering bags
![Page 57: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/57.jpg)
www.drsarma.in57
Diluting Cells
Transfer cells to Bag 2
(125mL wash media)
Rinse the CBU bag twice
with an additional 25 ml
of the Dextran/Albumin
solution from Bag 1.
Drain into Bag 2
The total volume in the
Bag 2 should be
approximately 200 mL
Bag 2
Bag 1
![Page 58: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/58.jpg)
www.drsarma.in58
First Centrifugation
Heat seal and remove
the CBU and Bag 1
(Dextran/Alb)
Centrifuge cells and
remaining bag @ 880
x g for 20 minutes at
4C.
![Page 59: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/59.jpg)
www.drsarma.in59
Expressing Supernatant
Express the
supernatant into the
attached transfer bag
Leave approx. 25 mL
of cell suspension in
bag.
Heat seal and
disconnect CBU from
supernatant bag
![Page 60: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/60.jpg)
www.drsarma.in60
Spinning the Supernatant
Connect a 300 mL transfer
bag to the supernatant
bag.
Centrifuge the supernatant
bag and attached transfer
bag to 880 x g for 15
minutes at 4C.
Express the supernatant
leaving approximately 10-
20 mL of cell suspension in
the original bag.
![Page 61: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/61.jpg)
www.drsarma.in61
Final Pooling
Combine cells
recovered from the 1st
and 2nd spins into 1
bag.
Add additional
Dextran/Albumin
solution so that final
product is 50mL (peds)
and 100mL (adults)
![Page 62: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/62.jpg)
www.drsarma.in62
QC Testing
QC Samples
Nucleated Cell
Hematocrit
Viability
ABO/Rh
CFU
CD34
Sterility
![Page 63: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/63.jpg)
www.drsarma.in63
Checking the Cord Blood for Disease
Syphilis
HIV Ag and Ab
Hepatitis B and C
ALT
Malaria
Leptospirosis
HTLV 1 and 2, CMV, EB Virus
HLA typing is not done routinely while banking. It is done only before Stem Cell transplant
![Page 64: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/64.jpg)
www.drsarma.in64
Final Product
![Page 65: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/65.jpg)
www.drsarma.in65
Potential Safety Issues
Older stem cells can turn cancerous, e.g.Human adipose stem cells in animalsUnexpected in adult stem cellsProblem is a function of age (number
of divisions outside body)Cell cultures should be no more than
60 generations old Need for pre-clinical research prior to
therapeutic trials
![Page 66: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/66.jpg)
www.drsarma.in66
International regulations regarding embryonic stem cell research A: Prohibition B: Utilisation C: Utilisation and
extraction D: Utilisation, extraction and creation
Austria Brazil Costa Rica Denmark Equador Ireland Peru Poland
Germany USA
Australia Canada Greece Japan Spain Sweden The Netherlands France
Singapore UK Israel China
[Taken from Towns C.R. Embryonic Stem Cell Research: Understanding and Analysis at the Interfaces of Science, Medicine and Ethics. PhD Thesis, University of Otago, 2004]
![Page 67: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/67.jpg)
www.drsarma.in67
![Page 68: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/68.jpg)
www.drsarma.in68
![Page 69: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/69.jpg)
www.drsarma.in69
American Association for Advancement of Science
![Page 70: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/70.jpg)
www.drsarma.in70
To learn more about theStem Cell Research visit
www.stemcells.nih.gov
National Marrow Donor Program (NMDP) www.marrow.org
This presentation is on the our website
www.drsarma.in
![Page 71: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/71.jpg)
www.drsarma.in71
International Stem Cell Forum
http://stemcellforum.org
Currently representatives from the following countries:
• Australia
• Canada
• Czech Republic
• Finland
• France
• Germany
• Israel
• Japan
• Singapore
• Sweden
• Switzerland
• The Netherlands
• UK
• USA (NIH)
JDRF is the only non-government partner
JDRF funds outside national borders
![Page 72: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/72.jpg)
www.drsarma.in72
Cord Blood Banking Facilities
Public Cord Blood BankingReliance Life Science (RLS), Bombay
2001– RLS has net work all over India 2005– ReliCord-S and ReliCord-A
Histostem ( of Korea) bank in Mumbai,– Additional centres for Delhi, Chennai
and Kolkata. – Govt of India has 10% equity stake
Private Cord Blood Banking – 2004 byLifeCell of ACCPL (Asia Cryo Cell P ltd)
– Affiliate of Cryo-Cell International (USA)
![Page 73: Dr R V S N Sarma MD MSc Consultant Physician & Chest Specialist](https://reader036.vdocuments.mx/reader036/viewer/2022081800/56815a79550346895dc7e0b2/html5/thumbnails/73.jpg)
www.drsarma.in73