ORAL REVALIDA

IH/BB, MBD, CC, HEMA, HISTOPATH​

BSMT 3-2 | PANGILINAN, BRYAN FROZ R. | SEM 2, 2024-2025

IH/BB TOPICS
Example:

●​ ABO Discrepancies (causes and Forward Typing Reverse

remedy) Typing
●​ Donor Screening (allogeneic
donation requirements, Anti-A: 0 A cells: 4+
collection and deferral)
●​ Hepatitis B Profile (HbsAg, Anti-B: 0 B cells: 4+
Anti-Hbs, HbeAg, Anti-HBc IgM,
Anti-HBc IgG, Anti-Hbe, HBV ●​ Interpretation: Apparent group
DNA) O, but reverse grouping suggests
missing anti-A/B (e.g., elderly
patient).
ABO DISCREPANCIES
●​ Remedies:
CAUSES & REMEDIES
○​ Incubate serum with
reagent A/B cells at room
●​ ABO discrepancies occur when
temperature (RT) for
forward grouping (antigen
15–30 minutes or 4°C for
detection on RBCs) and reverse
15 minutes.
grouping (antibody detection in
○​ Add extra serum to
serum) results conflict.
enhance weak reactions.
●​ These discrepancies arise from
○​ Test with O cells and
technical errors, biological
autocontrol to rule out
variations, or pathological
non-specific reactions.
conditions.
●​ Discrepancies are categorized
into four groups:
Group II: Weak/Missing Antigens in
Forward Grouping

Group I: Weak/Missing Antibodies in

Reverse Grouping ●​ ABO subgroups (e.g., A₃, Aₓ, B₃)
with weakly expressed antigens.

●​ Newborns: Lack ABO antibodies ●​ Leukemia/Hodgkin’s disease:

(develop after 3–6 months). Reduced antigen expression.

●​ Elderly patients: Reduced ●​ Acquired B phenomenon:

antibody titers. Bacterial enzymes (e.g., E. coli)
modify A antigens to mimic B.
●​ Immunosuppressed individuals
(e.g., leukemia, chemotherapy,
bone marrow transplants).

●​ Hypogammaglobulinemia or
congenital immunodeficiency.

1
ORAL REVALIDA
IH/BB, MBD, CC, HEMA, HISTOPATH​
BSMT 3-2 | PANGILINAN, BRYAN FROZ R. | SEM 2, 2024-2025

Example: Example:

Forward Typing Reverse Forward Typing Reverse

Typing Typing

Anti-A: 4+ A cells: 0 Anti-A: 4+ A cells: 2+

Anti-B: 2+ B cells: 4+ Anti-B: 2+ B cells: 2+

●​ Interpretation: True group A ●​ Interpretation: False

with "acquired B" antigen. agglutination due to rouleaux.

●​ Remedies: ●​ Remedies:
○​ Acidify anti-B reagent ○​ Wash RBCs 6–8 times
(pH <6) – true B antigens with saline to remove
react; acquired B does plasma proteins.
not. ○​ Add saline dilution to
○​ Treat RBCs with acetic disperse rouleaux.
anhydride to reverse ○​ Use albumin replacement
bacterial modification. technique.
○​ Use enzyme-treated ○​
RBCs (e.g., papain) to
enhance weak antigen
detection. Group IV: Miscellaneous Causes

●​ These discrepancies between

Group III: Plasma/Protein
forward and reverse groupings
Abnormalities
are due to miscellaneous
problems.
●​ These discrepancies between ○​ Cold autoantibodies (e.g.,
forward and reverse groupings anti-I) causing
are caused by protein or plasma non-specific
abnormalities and result in agglutination.
rouleaux formation or ○​ Unexpected
pseudoagglutination. alloantibodies (e.g.,
○​ Rouleaux formation due anti-M, anti-Lea).
to elevated globulins (e.g., ○​ Cis-AB phenotype: Rare
multiple myeloma) or inheritance of A and B
fibrinogen. genes on one
○​ Plasma expanders (e.g., chromosome.
dextran) causing ○​ Bombay phenotype (Oh):
pseudoagglutination. Lack of H antigen; serum
○​ Wharton’s jelly in cord contains anti-A, anti-B,
blood samples. and anti-H.

2
ORAL REVALIDA
IH/BB, MBD, CC, HEMA, HISTOPATH​
BSMT 3-2 | PANGILINAN, BRYAN FROZ R. | SEM 2, 2024-2025

Example:
Group II Use
enzyme-treated
Forward Typing Reverse cells, acidify
Typing anti-B.
Anti-A: 0 A cells: 4+ Group III Wash RBCs, saline
dilution.
Anti-B: 0 B cells: 4+
Group IV Warm samples,
●​ Interpretation: Oh phenotype (no use absorption
H antigen). techniques.

●​ Remedies:
○​ Wash RBCs 6–8 times
with saline to remove DONOR SCREENING
plasma proteins. ALLOGENEIC DONATION
○​ Add saline dilution to REQUIREMENTS, COLLECTION AND
disperse rouleaux. DEFERRAL
○​ Use albumin replacement
technique.
REQUIREMENTS
●​ Age:
NOTES ○​ Minimum age: 16 years
(16-17 years require
●​ General Technical Errors: parental consent; donors
○​ Clerical Errors (mislabeled >65 years need physician’s
samples). approval).
○​ Incorrect cell suspension
concentration.
○​ Contaminated reagents or ●​ Weight:
uncalibrated centrifuges. ○​ At least 50 kg to donate
●​ Prevention: 450 mL of blood.
○​ Follow strict SOPs. ○​ Formula for adjusted
○​ Use controls (e.g., O cells, blood volume:
autocontrol). ■​ Volume to collect =
(Donor’s Weight
[kg] / 50) x 450
mL.
SUMMARY
●​ Hemoglobin/Hematocrit:
Discrepancy Type Key Resolution ○​ Hemoglobin:
Step ■​ ≥12.5 g/dL
(female).
Group I Incubate serum
■​ ≥13.0 g/dL (male).
longer, add more
○​ Hematocrit:
plasma.
■​ ≥38% (female).
■​ ≥39% (male).

3
ORAL REVALIDA
IH/BB, MBD, CC, HEMA, HISTOPATH​
BSMT 3-2 | PANGILINAN, BRYAN FROZ R. | SEM 2, 2024-2025

●​ Health Screening: ○​ Former residents of

○​ Normal VS: malaria-endemic areas (>6
■​ Pulse: 50-100 bpm. months stay)
■​ BP: Systolic
90-180, Diastolic ●​ 6 Months:
50-100 mmHg. ○​ Travel to malaria-endemic
○​ No signs of infection or areas (<6 months stay).
high risk behaviors. ○​ Avodart (dutasteride) use.

●​ 3 Years:
COLLECTION
○​ Soriatane (acitretin) use..
●​ Pre-Donation: ○​ Malaria Infection
○​ Registration: (asymptomatic
■​ Verify ID post-treatment).
(first-time donors)
and link to ●​ 1 Month:
previous records ○​ Proscar/Propecia/​
(repeat donors). Accutane use.
○​ Consent: Provide written
materials explaining risks ●​ 2 Days:
and eligibility. ○​ Aspirin use.

●​ Procedure: ●​ Pregnancy/Childbirth:
○​ Cleanse: Use PVP iodine ○​ 6 weeks post-abortion; 9
(4 cm area, scrubbed for months postpartum.
30 seconds).
○​ Needle: 16-17 gauge for ●​ Permanent Deferrals:
single venipuncture; ○​ HIV/HBV/HCV positive.
collect 450 ±45 mL within ○​ IV drug use or
15 minutes. needle-sharing.
○​ Protozoan diseases
●​ Post-Donation: (Chagas, Babesiosis).
○​ Apply pressure for 5 ○​ Cancer (except minor skin
minutes; monitor for cancer/cervical
reactions. carcinoma in situ).
○​ Provide refreshments and ○​ Creutzfeldt-Jakob disease
observe donor for 15 exposure.
minutes.

POST-DONATION TESTING
DEFERRAL CRITERIA
●​ Mandatory Testing:
●​ 12 Months: ○​ ABO/Rh typing.
○​ Sexual contact with HIV+ ○​ Infectious Markers:
individuals. ■​ HBsAg (repeat in
○​ Mucous membrane duplicate +
exposure to neutralization if
blood/non-sterile positive).
needles.

4
ORAL REVALIDA
IH/BB, MBD, CC, HEMA, HISTOPATH​
BSMT 3-2 | PANGILINAN, BRYAN FROZ R. | SEM 2, 2024-2025

■​ Anti-HBc,
HEPATITIS B PROFILE
Anti-HCV,
Anti-HIV ½ HBSAG, ANTI-HBS, HBEAG, ANTI-HBC
(confirm w/ IGM, ANTI-HBC IGG, ANTI-HBE, HBV
Western Blot). DNA
■​ Syphilis
(screening: ●​ The Hepatitis B Profile is a
RPR/VDRL; critical serological panel used to
confirmatory: diagnose, monitor, and manage
FTA-ABS). Hepatitis B Virus (HBV)
●​ Disposal: Quarantine and dispose infections.
if tests are abnormal.
●​ It includes seven key markers:
HBsAg, Anti-HBs, HBeAg,
DONOR REACTIONS &
Anti-HBe, Anti-HBc IgM,
MANAGEMENT
Anti-HBc IgG, and HBV DNA.
●​ Mild:
○​ Syncope (fainting): ●​ Each marker provides distinct
Remove needle, elevate insights into the phase of
legs, apply cold compress. infection, immune status, and
○​ Nausea: Slow breathing, treatment efficacy.
cold compress, provide
receptacle.
MARKER DEFINITIONS & CLINICAL
●​ Moderate:
SIGNIFICANCE
○​ Loss of consciousness:
Administer 95% O2 + 5%
CO2, monitor vitals. ●​ HBsAg (Hepatitis B Surface
●​ Severe: Antigen)
○​ Convulsions/Cardiac ○​ Role: Indicates current
arrest: Restrain donor, HBV infection (acute or
perform CPR, call chronic).
physician. ○​ Clinical Insight: Detected
during the incubation
period and acute phase.
NOTES Persistence >6 months
●​ Record retention: suggests chronic
○​ 5-10 years for donor records. infection.
○​ Indefinite for notification of
abnormal results. ●​ Anti-HBs (Antibody to HBsAg)
●​ Deferrals: Protect donors (e.g., low
hemoglobin → anemia risk) and
○​ Role: Indicates immunity
recipients (e.g., transfusion-transmitted (from vaccination or
infections). resolved infection).
●​ Testing: Ensures blood safety (e.g., NAT ○​ Clinical Insight: A titer
detects early HIV/HCV).
>10 mIU/mL
●​ Reactions: Protocols minimize harm
(e.g., syncope management prevents post-vaccination confirms
injury). immunity.

5
ORAL REVALIDA
IH/BB, MBD, CC, HEMA, HISTOPATH​
BSMT 3-2 | PANGILINAN, BRYAN FROZ R. | SEM 2, 2024-2025

○​ Clinical Insight: High

●​ HBeAg (Hepatitis B e Antigen) levels (>20,000 IU/mL)
○​ Role: Reflects active viral indicate active
replication and high replication. Used to guide
infectivity. antiviral therapy.
○​ Clinical Insight: Presence
correlates with high HBV
INTERPRETATION OF KEY PROFILES
DNA levels.
Disappearance
(seroconversion to ●​ Acute HBV Infection:
Anti-HBe) often signals ○​ HBsAg (+)
reduced viral activity. ○​ Anti-HBc IgM (+)
○​ HBeAg (+).
●​ Anti-HBe (Antibody to HBeAg)
○​ Role: Indicates ●​ Chronic HBV Infection:
seroconversion and ○​ HBsAg (+) >6 months
reduced infectivity. ○​ Anti-HBc IgG (+)
○​ Clinical Insight: Appears ○​ HBeAg (+/-).
after HBeAg clearance. In
chronic infection, it may ●​ Resolved Infection:
signal transition to an ○​ Anti-HBc IgG (+)
inactive carrier state. ○​ Anti-HBs (+).

●​ Anti-HBc IgM (IgM Antibody to ●​ Vaccination Immunity:

Hepatitis B Core Antigen) ○​ Anti-HBs (+) alone (no
○​ Role: Marker of acute Anti-HBc).
infection (first antibody
to appear). DIAGNOSTIC METHODS &
○​ Clinical Insight: Elevated CONSIDERATIONS
in acute HBV; absent in
chronic cases (unless
reactivation). ●​ Screening:
○​ ELISA/Chemiluminescent
●​ Anti-HBc IgG (IgG Antibody to Immunoassay (automated,
Hepatitis B Core Antigen) batch testing).
○​ Role: Indicates past or
ongoing infection. ●​ Confirmation:
○​ Clinical Insight: Persists ○​ HBV DNA PCR for viral
for life. Differentiates load; HBsAg neutralization
resolved infection tests to rule out false
(Anti-HBc IgG + Anti-HBs) positives.
from vaccination
(Anti-HBs alone). ●​ False Positives:
○​ Repeat testing and
●​ HBV DNA molecular confirmation
○​ Role: Quantifies viral load required.
and assesses treatment
response.

6
ORAL REVALIDA
IH/BB, MBD, CC, HEMA, HISTOPATH​
BSMT 3-2 | PANGILINAN, BRYAN FROZ R. | SEM 2, 2024-2025

○​ DNA ligase seals nicks

MBD
between Okazaki
fragments.
●​ DNA replication is the
semi-conservative process
(Meselson & Stahl, 1958) by which NOTES
DNA synthesizes a new
●​ Enzymes & Components:
complementary strand using the
○​ DNA helicase: Unwinds
original parent strand as a
DNA.
template.
○​ DNA polymerase:
Synthesizes new strands (I
for primer removal, III for
DNA REPLICATION elongation).
EXPLANATION ○​ Primase: Creates RNA
primers.
●​ Initiation: ○​ Ligase: Joins DNA
○​ Origin of replication: fragments.
Specific DNA sequences ●​ Significance:
where replication begins. ○​ Essential for cell division,
○​ DNA helicase unwinds the growth, and tissue repair.
double helix, creating a ○​ Ensures genetic fidelity
replication fork. through
○​ Single-strand binding semi-conservative
proteins (SSBs) stabilize replication (each
unwound DNA. daughter DNA has one
parent strand).
●​ Elongation:
○​ DNA primase synthesizes
short RNA primers.
○​ DNA polymerase III adds PROCESS OF PCR
nucleotides to the 3’ end
of the primer, forming EXPLANATION
new strands:
■​ Leading strand: ●​ PCR, developed by Kary Mullis
Synthesized (1983), is a laboratory technique
continuously. to amplify specific DNA
■​ Lagging strand: sequences exponentially.
Synthesized
discontinuously as ●​ Components of PCR:
Okazaki fragments. ○​ Template DNA: Contains
the target sequence to be
●​ Termination: amplified (e.g., genomic
○​ DNA polymerase I DNA, plasmid DNA).
replaces RNA primers ○​ Primers: Short, synthetic
with DNA. oligonucleotides (18–25
bp) that flank the target
region.

7
ORAL REVALIDA
IH/BB, MBD, CC, HEMA, HISTOPATH​
BSMT 3-2 | PANGILINAN, BRYAN FROZ R. | SEM 2, 2024-2025

■​ Forward and DNA strands from

Reverse Primers: primers.
Define the start
and end of the ●​ Cycle Progression:
amplified ○​ Exponential
sequence. Amplification: Each cycle
○​ Taq Polymerase: doubles DNA (e.g., 30
Heat-resistant DNA cycles → ~1 billion copies).
polymerase from Thermus ○​ Plateau Phase: Reaction
aquaticus (optimal activity slows as reagents deplete.
at 72°C).
○​ dNTPs: Deoxynucleotide
TYPES OF PCR
triphosphates (A, T, C, G)
for DNA synthesis.
○​ Buffer Solution: ●​ Reverse Transcription PCR
Maintains pH (e.g., (RT-PCR):
Tris-HCl) and provides ○​ Converts RNA to cDNA
Mg²⁺ (cofactor for Taq). using reverse
transcriptase before
●​ Thermal Cycling Steps: amplification.
○​ Denaturation: ○​ Application: Detecting
■​ Temperature: RNA viruses (e.g.,
94–98°C for 20–30 SARS-CoV-2).
seconds.
■​ Action: Separates ●​ Quantitative PCR (qPCR):
double-stranded ○​ Measures DNA
DNA into single accumulation in real-time
strands by using fluorescent dyes
breaking hydrogen (e.g., SYBR Green) or
bonds. probes (TaqMan).
○​ Annealing: ○​ Application: Quantifying
■​ Temperature: viral load (e.g., HIV,
50–65°C hepatitis).
(primer-specific)
for 20–40 seconds. ●​ Multiplex PCR:
■​ Action: Primers ○​ Amplifies multiple
bind to targets simultaneously
complementary using multiple primer
sequences on pairs.
template DNA. ○​ Application: Pathogen
○​ Extension: panels (e.g., respiratory
■​ Temperature: 72°C viruses).
for 1–2 minutes
(depending on ●​ Nested PCR:
target length). ○​ Uses two rounds of PCR
■​ Action: Taq with nested primers to
polymerase improve specificity.
synthesizes new

8
ORAL REVALIDA
IH/BB, MBD, CC, HEMA, HISTOPATH​
BSMT 3-2 | PANGILINAN, BRYAN FROZ R. | SEM 2, 2024-2025

○​ Application: Detecting ●​ Research:

low-abundance targets ○​ Cloning: Amplifies genes
(e.g., Mycobacterium for insertion into
tuberculosis). plasmids.

PCR OPTIMIZATION & CLINICAL CHEMISTRY

TROUBLESHOOTING
●​ End product of hemoglobin
Issue Cause Solution metabolism and the principal
pigment of bile
No - Primer - Redesign primers ●​ Orange yellow pigment derived
Product mismatch - Adjust Mg²⁺ from hemoglobin degradation
- Low Mg²⁺
●​ Also formed from destruction of
Non-Sp - High primer - Titrate primers heme-containing proteins such
ecific concentratio - Increase as myoglobin, catalase and
Bands n annealing temp cytochrome oxidase.
- Low
annealing
temp BILIRUBIN METABOLISM (PATHWAY)
Primer - - Improve primer
Dimers Self-comple design (e.g., check ●​ Sources:
mentary GC content)
○​ Hemoglobin (75-80%):
primers
From senescent RBCs
Contam - Foreign - Use UV hoods broken down by
ination DNA (e.g., lab - Separate pre- macrophages in the
aerosols) and post-PCR
spleen and liver.
areas
○​ Heme proteins (20-25%):
Myoglobin, cytochromes,
APPLICATION IN MOLECULAR catalase.
DIAGNOSTICS
●​ Pathway Steps:
○​ Degradation of Heme:
●​ Infectious Diseases:
■​ Macrophages
○​ COVID-19: RT-PCR
convert heme to
detects viral RNA in
biliverdin via heme
nasopharyngeal swabs.
oxygenase.
○​ HIV: qPCR monitors viral
■​ Biliverdin is
load during antiretroviral
reduced to
therapy.
unconjugated
●​ Genetic Disorders:
bilirubin (indirect,
○​ BRCA1/2 Mutations: PCR
lipid-soluble).
amplifies exons for
sequencing to assess
○​ Transport to Liver:
breast cancer risk.
■​ Unconjugated
●​ Forensics:
bilirubin binds to
○​ STR Analysis: Amplifies
albumin (prevents
short tandem repeats for
DNA fingerprinting.

9
ORAL REVALIDA
IH/BB, MBD, CC, HEMA, HISTOPATH​
BSMT 3-2 | PANGILINAN, BRYAN FROZ R. | SEM 2, 2024-2025

renal excretion due

MECHANISM OF RAAS
to insolubility).
○​ Hepatic Uptake:
■​ Hepatocytes take ●​ Trigger: ↓ → ↑
up bilirubin via ○​ ↓ Blood flow to kidneys
ligandin (binding (e.g., dehydration,
protein). hemorrhage, heart
failure).
○​ Conjugation: ○​ Specialized kidney cells
■​ UDP-glucuronyl (juxtaglomerular cells)
transferase in detect this drop and
hepatocytes adds release renin.
glucuronic acid →
conjugated ●​ Renin → Angiotensin I:
bilirubin (direct, ○​ Renin (an enzyme) acts on
water-soluble). angiotensinogen (a
liver-made protein).
○​ Biliary Excretion: ○​ Renin then cuts or cleaves
■​ Conjugated angiotensinogen into
bilirubin is angiotensin I (inactive
excreted into bile peptide).
→ small intestine.
●​ Angiotensin I → Angiotensin II
○​ Intestinal Metabolism: via ACE:
■​ Gut bacteria ○​ ACE
convert conjugated (Angiotensin-Converting
bilirubin to Enzyme) in the lungs
urobilinogen: converts angiotensin I →
■​ Stercobilin angiotensin II (active
(oxidized to brown hormone).
pigment in feces).
■​ Urobilin ●​ Angiotensin II actions:
(reabsorbed → ○​ Vasoconstriction:
enters Squeezes blood vessels →
enterohepatic ↑ blood pressure.
circulation; 20% ○​ Stimulates Aldosterone
excreted in urine). Release: Signals adrenal
glands to release
aldosterone.
○​ Triggers Thirst: Acts on
the brain to increase
water intake.
○​ ADH Release: Promotes
water retention in
kidneys.

10
ORAL REVALIDA
IH/BB, MBD, CC, HEMA, HISTOPATH​
BSMT 3-2 | PANGILINAN, BRYAN FROZ R. | SEM 2, 2024-2025

●​ Aldosterone Role: Complications Microvascular Macrovascular

○​ Aldosterone acts on (retinopathy, (CVD, stroke),
nephropathy) neuropathy
kidney tubules to:
■​ Retain sodium →
Water follows ●​ Diagnostic Criteria:
sodium → ↑ blood ○​ HbA1c ≥6.5%
volume. (NGSP-certified method).
■​ Excrete potassium ○​ Fasting Plasma Glucose
→ Maintains ≥126 mg/dL.
electrolyte ○​ 2-hr OGTT ≥200 mg/dL.
balance. ○​ Random Glucose ≥200
mg/dL + symptoms
(polyuria, polydipsia).
DIABETES MELLITUS: TYPE I vs TYPE
II
●​ Pathophysiology
○​ Type 1: Absolute insulin
●​ Diabetes mellitus is actually a deficiency → ↑ lipolysis →
group of metabolic diseases ketogenesis (DKA).
characterized by hyperglycemia ○​ Type 2: Insulin resistance
resulting from defects in insulin → ↑ hepatic glucose
secretion, insulin action, or both. output + ↓ peripheral
uptake → hyperglycemia.
Feature Type 1 Type 2
Diabetes Diabetes ●​ Lab Findings:
○​ DKA (Type 1):
Pathogenesis Autoimmune Insulin
β-cell resistance + ■​ ↑ Glucose (>250
destruction progressive mg/dL)
(anti-GAD, IA-2 β-cell
antibodies) dysfunction
■​ ↑
β-hydroxybutyrate
Onset Acute (weeks), Insidious, ■​ ↓ pH (<7.3)
often <30 years typically >40
years
■​ Anion gap >12.
○​ Hyperosmolar State
Body Weight Normal or Overweight/o (Type 2):
underweight bese (80% of
cases)
■​ ↑ Glucose (>600
mg/dL)
Ketosis Common (DKA) Rare (unless ■​ ↑ Osmolality (>320
stressed;
hyperosmolar
mOsm/kg)
coma) ■​ No ketosis.

Insulin Absent Normal or ↑

Secretion early; ↓ late
●​ Management:
○​ Type 1: Basal-bolus insulin
Autoantibodies Present Absent regimens, CGM
(GAD65, ICA,
IA-2) (continuous glucose
monitoring).
Treatment Mandatory Lifestyle, oral ○​ Type 2: Metformin
insulin agents
(metformin), ± (first-line), SGLT2
insulin inhibitors, GLP-1 agonists.

11
ORAL REVALIDA
IH/BB, MBD, CC, HEMA, HISTOPATH​
BSMT 3-2 | PANGILINAN, BRYAN FROZ R. | SEM 2, 2024-2025

●​ Special Cases: vasoconstriction and

○​ LADA (Latent recruits platelets.
Autoimmune Diabetes):
Slow-onset Type 1 in ●​ Aggregation:
adults. ○​ GPIIb/IIIa binds
○​ MODY (Maturity-Onset fibrinogen, forming a
Diabetes of the Young): temporary plug.
Monogenic form (e.g.,
HNF1A mutations). ●​ Clinical Relevance:
○​ Deficiencies cause
bleeding disorders (e.g.,
NOTES
Bernard-Soulier
●​ Bilirubin: Conjugation defects vs. syndrome, Glanzmann
excretion disorders determine thrombasthenia).
jaundice type.
MECHANISM OF HEMOSTASIS
●​ RAAS: Central to BP/volume
SECONDARY HEMOSTASIS
regulation; targeted in
hypertension/heart failure.
●​ Extrinsic Pathway:
○​ TF activates FVIIa,
●​ Diabetes: Autoimmunity vs.
initiating coagulation.
metabolic syndrome dictate
management.
●​ Intrinsic Pathway:
○​ Contact activation (FXII,
HEMATOLOGY collagen) amplifies via
FXI, FIX, FVIII.

MECHANISM OF HEMOSTASIS ●​ Common Pathway:

○​ FXa converts
PRIMARY HEMOSTASIS
prothrombin to
thrombin, which cleaves
●​ Vascular Injury:
fibrinogen to fibrin. FXIIIa
○​ Endothelial damage
cross-links fibrin.
exposes subendothelial
collagen.
●​ Regulation:
○​ Antithrombin, protein
●​ Platelet Adhesion:
C/S, and TFPI localize
○​ VWF binds collagen and
clotting. Thrombin binds
platelets via GPIb/IX
thrombomodulin to
(critical under high shear
activate protein C
stress).
(inhibits FVa/VIIIa).
●​ Platelet Activation:
○​ Release of Adenosine
diphosphate (ADP),
Thromboxane A2 (TXA2),
and serotonin induces

12
ORAL REVALIDA
IH/BB, MBD, CC, HEMA, HISTOPATH​
BSMT 3-2 | PANGILINAN, BRYAN FROZ R. | SEM 2, 2024-2025

MECHANISM OF HEMOSTASIS
●​ Dehydration Issues
FIBRINOLYSIS ○​ Incomplete Dehydration:
Results in residual water,
●​ Clot Breakdown: tPA (from preventing proper
endothelium) activates paraffin infiltration → soft
plasminogen → plasmin blocks.
degrades fibrin into D-dimers. ○​ Over-Dehydration: Makes
tissues brittle, leading to
●​ Inhibition: PAI-1 and fragmentation during
α2-antiplasmin prevent sectioning.
excessive lysis. TAFI stabilizes ●​ Clearing Problems
clots by removing lysine ○​ Inadequate Clearing:
residues. Residual dehydrating
agent (e.g., alcohol)
MECHANISM OF HEMOSTASIS hinders paraffin
infiltration.
CLINICAL TESTING ○​ Prolonged Clearing:
Overexposure to
●​ Bleeding Time: Assesses platelet xylene/toluene makes
function. tissues hard and crumbly.

●​ PT/INR: Extrinsic pathway. ●​ Infiltration and Embedding

○​ Poor Paraffin Infiltration:
●​ APTT: Intrinsic pathway. Causes uneven
embedding, resulting in
●​ D-dimer: Fibrinolysis marker holes or tears in sections.
(elevated in DIC, VTE). ○​ Improper Orientation:
Tissue misalignment in
HISTOPATH the block complicates
sectioning.
○​ Air Bubbles: Trapped
DIFFICULTIES AND ERRORS IN TISSUE bubbles distort tissue
PROCESSING AND SECTIONING architecture.
TISSUE PROCESSING
DIFFICULTIES AND ERRORS IN TISSUE
●​ Fixation Errors PROCESSING AND SECTIONING
○​ Incomplete Fixation: SECTIONING
Leads to poor tissue
preservation, autolysis, or
●​ Block Preparation
artifactual changes.
○​ Excessive Trimming:
○​ Over-Fixation: Causes
Removes too much tissue,
excessive hardening,
risking loss of diagnostic
making sectioning
material.
difficult.
○​ Insufficient Trimming:
Leaves excess wax,

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ORAL REVALIDA
IH/BB, MBD, CC, HEMA, HISTOPATH​
BSMT 3-2 | PANGILINAN, BRYAN FROZ R. | SEM 2, 2024-2025

hindering ribbon slides if not properly

formation. dried.

●​ Microtome Issues
HISTOPATH
○​ Dull or Damaged Knives:
Cause tearing, chatter
(horizontal lines), or ●​ Equipment Maintenance
thick-thin sections. ○​ Regularly hone/strop
○​ Incorrect Knife Angle: knives; replace disposable
Poor clearance/rake blades.
angles lead to uneven cuts ○​ Clean microtome with
or knife marks. xylol and oil moving parts
○​ Loose Microtome Parts: to prevent rust.
Vibrations create irregular
sections. ●​ Optimal Conditions
○​ Maintain paraffin
●​ Section Quality embedding temperature
○​ Chatter: Repetitive (2–3°C below melting
thickness variations due point).
to vibration or dull blades. ○​ Set water bath 6–10°C
○​ Wrinkles/Folds: Caused below paraffin melting
by improper water bath point (typically 45–50°C).
temperature or rapid
section expansion. ●​ Technique Adjustments
○​ Tears/Cracks: Brittle ○​ Trim blocks cautiously:
blocks (over-dehydration) Coarse trimming (30 µm)
or hard tissues (e.g., bone). → fine trimming (15 µm).
○​ Use appropriate knife
●​ Floating and Adhesion profiles (e.g., wedge knife
○​ Water Bath Temperature: for hard tissues).
Too hot → melts paraffin; ○​ Adjust microtome settings
too cold → sections fail to (clearance angle: 5–10°;
flatten. section thickness: 4–6 µm
○​ Bubbles Under Sections: for paraffin).
Trapped air distorts tissue
morphology. ●​ Troubleshooting
○​ Overexpansion: ○​ Brittle Blocks: Rehydrate
Prolonged floating (>30 or use a softer wax
seconds) stretches ○​ Chatter: Tighten
tissues. microtome, sharpen
knives, or reduce section
●​ Freezing Microtome/Cryostat thickness.
Challenges ○​ Wrinkles: Adjust water
○​ Ice Crystal Formation: bath temperature or
Poor freezing technique gently tease sections with
damages cellular details. a brush.
○​ Tissue Adhesion Issues:
Sections may detach from

14