Nutr. 112 - LAB 5
2014
Week 5 LAB: Folate and vitamin B12 Assessment
Background and Overview
 Deficiencies of either Folate and vitamin B12 can result in megaloblastic anemia - an anemia
characterized by the presence of abnormally large, immature red blood cell precursors (megaloblasts)
in the bone marrow, and an increased proportion of larger than normal (macrocytic), immature RBCs
(reticulocytes) in the peripheral blood.
 Because of the role and importance of Folate in neonatal development, many industrialized countries
now fortify cereal grain products (including flour, rice, breads, pasta, noodles and corn meal) with
Folate. This includes the US, which began mandatory fortification in 1998.
 B12 deficiency can also result in nerve disorders and spinal cord dysfunction- even in the absence of
anemia. These neurological effects typically take years of B12 deficiency to manifest, and can cause
both reversible and irreversible damage.
 Folate deficiency generally develops more quickly than B12 deficiency because of the differences in
turnover and utilization rates of the two nutrients. Inadequate intake is the major factor underlying
folate deficiency, while B12 deficiency generally arises for reasons relating to absorption, including
the inadequate production or secretion of intrinsic factor.
 Both Folate and vitamin B12 serve as coenzymes involved in a variety of biochemical pathways,
some of which are inter-connected.
Key metabolic/biochemical aspects:
 Folate and B12 are required for production of 5,10-methylene tetrahydrofolate, critical in the
synthesis of thymidine, and are thus both are essential for DNA synthesis and cell replication.
 Generation of the active, coenzyme form of Folate requires B12, while Folate serves as a methyl
donor to B12 in the conversion of homocysteine to methionine. Thus, in B12 deficiency, Folate
can become functionally “trapped” in a form that cannot be used metabolically.
Stages in the Progression to Deficiency
Most nutrient deficiencies occur in sequential stages:
Adequate  early depletion  depletion  early deficiency  deficiency (overt clinical signs – Anemia).
Identifying where a subject is in this progression is dependent on the presence of several distinct biomarkers
or indice (cut-off) values. Keep in mind that the values and transitions between stages may not be perfectly
clear cut. As we saw in earlier labs, the cutoff was based on optimizing sensitivity and specificity. The folate
deficiency sequence and cut-offs is provided in the appendix that follows.
Signs and symptoms of Folate and B12 Deficiency
The most prominent clinical sign of folate deficiency is the presence of macrocytic RBCs in the peripheral
blood. Should the deficiency persist over several months, an overt clinical megaloblastic anemia becomes
manifest. Although macrocytosis is a hallmark of either Folate or B12 deficiency, it should be noted that
there are a number of other conditions that can also result in the appearance of macrocytic RBCs.
CONDITION
Alcoholism
Liver disease
Renal disease
Hemolysis
Post-hemorrhagic Anemia
CAUSE of MACROCYTOSIS
Direct effect of on bone marrow. Impaired GI absorption
Defective DNA synthesis or  erythropoiesis
Decreased erythropoiesis ( production of erythropoietin)
 erythropoiesis
 erythropoiesis
Nutr. 112 - LAB 5
2014
Measurement of Folate
(1) Microbiological Assay – used for both folate and B12 assessment, wide-spread use
Utilizes a special strain of L. casei bacteria. Assay measures bacterial growth-rate in the presence of
serum/plasma or tissue extracts (with growth rate proportional to the folate concentration).
(2) Competitive Binding Assay – available to a limited extent
Utilizes a specific, high-affinity binding protein derived from milk, together with labeled folate. The
assay measures the amount of folate by calculating the quantity of radioactivity displaced (not bound) in
a sample. Assay is rapid and not generally effected by drugs or antibiotics which may otherwise interfere
with the microbiological assay.
(3) High Performance Liquid Chromatography (HPLC) – availability limited
Utilizes size, polarity and wavelength specific separation to directly measure the compound of interest.
Further modifications include Mass Spectroscopy as part of the analysis.
Assessment of Folate Status – Indices and Biomarkers
Serum (or Plasma) Folate
The predominant form of folate in the circulation (serum/plasma) is the reduced derivative,
methyltetrahydrofolate (mTHF). Serum/plasma folate concentrations generally fluctuate in response to
recent changes in folate intake. Thus, serum/plasma folate reflects primarily acute status.
Factors / Conditions which can result in INCREASED serum Folate
1) Renal Failure/dysfunction 2) Liver damage/disease 3) RBC hemolysis
4) B12 Deficiency (sometimes)
Factors / Conditions which can result in DECREASED serum Folate
1) Chronic alcohol ingestion 2) Chronic smoking / smoke exposure 3) Pregnancy (particularly late)
4) Certain drugs & medications (notably folate antagonists used in chemotherapy; some anti-inflammatory,
anti-asthma, or anti-convulsant drugs)
Erythrocyte (RBC) Folate
Erythrocytes take up and use folate as they develop in the bone marrow. Once in the circulation, uptake
ceases and levels remain relatively constant over the RBC’s lifespan. Thus, RBC folate is far less sensitive
than serum folate to short-term fluctuations, and reflects longer-term status and tissue stores.

RBC folate is effected by the same factors as for serum/plasma (noted above). However, of particular
note is the effect of vitamin B12 on RBC folate levels.
 vit. B12 =>  RBC Folate
Cellular folate uptake is B12 dependent.
Serum Homocysteine (tHcy)
 RBC Folate =>  Homocysteine
The favored biochemical pathway for catabolism of homocysteine in the fasted state, requires B12 as a
coenzyme and folate as a co-substrate (methyl donor). Thus, with deficiency of either folate or B12, Hcy
cannot be broken down as readily and accumulates.
Nutr. 112 - LAB 5
2014
Neutrophil Lobe Count (Lobe ave.)
 vit. B12 or  Folate => >3.5 lobes/cell
Neutrophils usually have only 3 or 4 nuclear segments (“lobes”). However, in megaloblastic anemia, the
number of these segments (lobes) increases. This hypersegmentation of neutrophils in peripheral blood may
often be the earliest morphological change to appear with folate or B12 deficiency; preceding the development
of macrocytosis. However, this can only be discerned in a blood smear analysis, which may (or may not) be
part of the CBC report. So, while a valuable, additional piece of data, it may not be available in all instances.
Also, there is no relationship between the lobe ave. and the severity of deficiency.
Deficiency of vitamin B12
Clinical signs of B12 deficiency occur in the hematopoietic (blood cell), gastrointestinal and neurological
systems. The hematological effects are indistinguishable from those of Folate deficiency. The gastrointestinal
effects of B12 deficiency include papillary atrophy of the tongue, loss of appetite and constipation. The
neurological complications are variable and progress gradually. These may precede or occur without anemia.
Manifestations include tingling and numbness of the extremities, motor disturbances and cognitive changes.
Neurological complications may become irreversible even after B12 treatment.
B12 deficiency is relatively rare, and most often results from malabsorption rather than overt dietary
deficiency (in developed countries). Lack of intrinsic factor, intestinal parasites, and alcoholism can result in
B12 deficiency.
Pernicious anemia is a multi-factorial disease with an autoimmune basis, characterized by an absence or
markedly reduced functional levels of gastric Intrinsic Factor (IF), a protein made by stomach parietal cells
that binds B12 and promotes its transport to the ileum for absorption. The majority of individuals with
pernicious anemia can be identified based on the presence of serum antibodies directed against either the IF
protein itself (type 1 IF-Ab), or to the ileal cell IF-receptor (type 2 IF-Ab). However, there are some
individuals with pernicious anemia who do not express either of these IF-Abs.
Deficiency of IF is often a consequence of the presence of atrophic gastritis, (a condition sometimes brought
on after a prolonged infection), which results in the destruction of parts of the stomach mucosa where parietal
cells are found. These individuals are positive for auto-antibodies against their own Parietal cells (PC-Abs).
Thus, diagnosis of pernicious anemia can be confirmed by identifying and measuring IF-Abs and PC-Abs
levels in the serum. Patients who are negative for IF-Abs should ideally undergo Schilling’s test to
definitively establish or refute the PA diagnosis. The onset and progression of PA is typically very slow
(often taking years). As a consequence, patients are often not aware of their symptoms – especially those
related to anemia, because over time they have become used to them.
Measurement of Vitamin B12 in Fluids and Tissues
As the case for Folate, competitive-binding assays (here using labeled B12) and microbiological assays (using
L. leichmannii or Euglena gracilis – distinct from the bacterial strains used to measure Folate) are technically
the same as employed for Folate. Here also, the semi-automated microbiological assay is still common,
although the use of radioimmunoassay kits for analysis of B12 is becoming increasingly widespread.
Assessment of vitamin B12 Status – Indices and Biomarkers
Total Serum B12
While total serum B12 is the biomarker most often measured, approximately 20% of the B12 in serum is
bound to transcobalamin (TC), with the remaining ~80% bound by haptocorrin (HC). Total serum B12 levels
Nutr. 112 - LAB 5
2014
have been observed to vary markedly in healthy persons, based in part on the measurement method. Thus,
while cut-offs have been used, total serum B12 is most valuable when used in combination with biomarkers.
Serum holoHC
holoHC is the major fraction of B12 in the circulation. Levels are typically not very responsive to recent
intake, thus it appears more aligned with longer-term status. Its measurement is not as common as either
total serum B12 or some of the other biomarkers.
RBC- B12
As with folate, the B12 in RBCs is a reflection of stores. However, this measurement is largely in the
developmental stage and not yet in common, wide-spread use.
Serum holoTC
Although a minor fraction in the circulation, holoTC is the physiologically “active” form of B12 delivered to
cells. Because holoTC has a short half-life, levels fall rapidly if B12 absorption ceases. Consequently, low
holoTC concentrations in serum are the earliest indicator of negative B12 balance (early depletion), falling
before total serum B12.
Serum Homocysteine (tHcy)
See above for Folate.
Methylmalonic Acid (MMA)
B12 serves as a cofactor for the enzyme that converts methylmalonyl CoA  succinyl CoA.
Because this reaction is independent of folate, MMA is highly specific for B12, and is not affected by Folate
deficiency. As B12 decreases, MMA accumulates in the blood (serum/plasma) and is then excreted in
increased amounts in the urine. MMA is more commonly measured in serum (versus urine), and is a
sensitive test for early functional B12 deficiency - before the onset of anemia. However, the MMA in does
not necessarily reflect the severity of the deficiency. And, since blood MMA levels can be affected by the
proportion eliminated in the urine, normal kidney function may have to be assessed.

As in folate assessment, no single biomarker provides conclusive B12 assessment data. Thus, assessment
of B12 is most effective using a combination of biomarkers/indices.
Evaluating vitamin B12 absorption
Once B12 deficiency has been diagnosed, it must still be determined whether it is a lack / low dietary intake or
inadequate B12 absorption, that is the problem. Distinguishing between these two potential root-causes is
approached using the tests below.
Schilling’s Test
The Schilling’s test measures B12 absorption of B12 - either in pure, crystalline form or bound with food (the
“Food Schilling’s Test”). The test is a 2-stage process requiring 2-3 days to perform. First, a small
radioactive dose (57Co-B12) is given, followed by a flushing dose of non-radioactive B12. Urine is then
collected over a 24 hr period and the amount of radioactivity in the urine is determined. In the second stage,
intrinsic factor (IF) is given along with a dose of radioactive B12 (an identical dose to that in stage 1). The
test will not be as indicative in situations of renal dysfunction, and is also be affected by certain GI
diseases/disorders and the presence of some intestinal parasites. Given the use of radioactivity and recent
issues in the manufacturing and availability of the test dose, the Schilling’s test is not used as much today.
Antibody Tests
Nutr. 112 - LAB 5
2014
Intrinsic Factor (IF) is an absolute requirement for B12 absorption. As noted above with respect to Pernicious
Anemia, a number of conditions can result in an autoimmune situation where the body produces antibodies
against one’s own proteins (auto-antibodies). Individuals who are positive for any of these auto-antibodies
(type 1 IF-Ab, type 2 IF-Ab, or ABG-Ab), will have impaired B12 absorption.
Nutr. 112 - LAB 5
2014
LAB 5 – APPENDIX
Biomarkers / Indices correlated with STAGES in DEFICIENCY PROGRESSION
Indice / Biomarker
NORMAL /
ADEQUATE
DEPLETION
EARLY
DEFICIENCY
DEFICIENCY
Serum Folate (nmol/L)
≥ 11.3
< 11.3
≤ 6.8
≤ 6.8
RBC Folate (nmol/L)
> 450
< 360
< 320
< 227
Neutrophil Lobe Ave.
< 3.5
< 3.5
≥ 3.5
≥ 3.5
Homocysteine (mol/L)
< 10
10 - 13
> 13
> 13
MMA (pmol/L)
< 210
210 - 270
> 270
> 270
Total ser B12 (pmol/L)
> 221
221 - 149
221 - 149
≤ 148
holoTC (pmol/L)
> 40
40 - 35
40 - 35
≤ 30
holoHC (pmol/L)
> 170
170 - 110
170 - 110
< 110
RDW (%)
< 14
≤ 14
> 14
> 14
MCV (fL)
80 - 100
80 - 100
80 - 100
> 100
outside normal
range
MCHC (g/dL)
32 - 36
32 - 36
May be outside
normal range
Hematocrit [ Hct ] (%)
normal
normal
normal
 or normal
Hemoglobin [ HgB ] (g/dL)
normal
normal
normal
 or normal
Values for Assessing RISK of DEFICIENCY
ACCEPTABLE
[ Adequate ]
BORDERLINE
[ MARGINAL ]
DEFICIENT
FOLATE biomarker
Serum Folate (nmol/L)
RBC Folate (nmol/L)
> 11.3
≥ 450
11.3 - 6.8
360 - 320
< 6.8
< 320
> 221
> 40
> 170
221 - 149
40 - 30
170 - 110
≤ 148
< 30
< 110
Vit. B12 biomarker
Total serum B12 (pmol/L)
holoTC (pmol/L)
holoHC (pmol/L)
MMA (Methylmalonic acid) (pmol/L)
> 270
RDW = (Std.Dev. MCV × 100) ÷ MCV
[ > 14 % = abnormal ]
MCV = Hct ÷ RBC conc. (× 1012 / L )
Normocytic = 80–100fL
MCH = [ HgB (g/dL) × 10 ] ÷ RBC conc. (× 1012 / L )
MCHC = HgB (g/dL) ÷ Hct (L/L)
or
Normochromic = 26–34 pg
MCH (pg) ÷ MCV (fL)
Normochromic = 32–36 g/dL (or %
2013 LAB 5
Name: _______________Problem Set _______________
Lab TA: ________________________
Week 5 - PROBLEM SET ( 243 points )
Note: Clinical values are reported to one decimal place; please report your final calculations accordingly.
Must show calculations and units for full credit.
(1) Folate/folic acid deficiency anemia can be characterized by: (2 points)
a)
b)
c)
d)
e)
High MCV and normal MCHC
normal MCV and low MCHC
Low MCH and high Hct
a and b only
none of the above
(2) Lab tests/indices and specific clinical terms are used to both define the presence of anemia as well as
provide morphological characteristics of blood cells which can then be used in ascribing or narrowing
down possible cause(s) of the anemia. Match the test/indice or clinical term listed at left to the
statements on the right. Note – each indice or term may have more than one letter match.
(23 points)
RDW
Poikilocytosis
Chromicity
Macrocytosis
MCHC
Hemoglobin (HgB)
Microcytosis
Anisocytosis
MCV
Hematocrit (Hct)
MCH
A. measure of the ave. size (ave. volume) of a RBC in the sample
B. indice may be significantly influenced by any change in cell size
C. measure of the concentration of hemoglobin in whole blood
D. a value used to establish clinical presence of anemia
E. would be influenced by significant hemolysis of a blood sample
F. value for the proportion of whole blood that are RBCs
G. will ultimately always be altered in anemia
H. indice for the ave. hemoglobin conc. per unit volume RBCs
I. measure of the size variation in a sample population of RBCs
J. indicates that there are abnormal gases within the blood sample
K. indice for the ave. HgB amount in an ave. RBC
L. term/value indicating the color intensity of RBCs
M. indicates there is variation in cell shape within a blood sample
N. indicates the ave. RBC is abnormally large (>100 fL)
O. is never observed or never changes with anemia
P. indicates the presence of different HgB sub-types in the RBCs
Q. refers to the presence of lysed cells within the blood sample
R. indicates the ave. RBC is abnormally small (<80 fL)
S. Term indicates an abnormal number of white blood cells
T. indicates that the ave. RBC is of normal size (80-100 fL)
U. indicates a variation in cell sizes within a blood sample
2013 LAB 5
Name: _______________Problem Set _______________
Lab TA: ________________________
(3) Just over 2 weeks ago (15 days ago), a 38 yr. old male subject presented with complaints of
“tiredness” and that he “fatigues easily” with any simple exertion. At that time a blood sample was
taken his CBC values were: RBCs = 3.94 x 1012 cells/L Hct = 41.6% HgB = 10.3 g/dL (25 points)
Based on these values, was he anemic? ( Yes / No ) (1 pt)
Based on: Low Hct / Low HgB / neither (both Normal) (1 pt)
(1 pt) Severity =
Using the values listed above, what were his values for: [ Show calculations, 6 points]
MCV =
MCH =
MCHC =
RBC Morphology (circle ALL that apply):
[ 1 pt.each ]
microcytic / normocytic / macrocytic
hypochromic / normochromic / hyperchromic
In reviewing his health history, you learn that he is a habitual 2 pack/day smoker. Which of the indices
could be affected by this? (Circle only those that could be affected) (5 points)
Hct
HgB
MCV
MCH
MCHC
All could be affected
He also reported that he started on a new diet about 2 months ago. Additional blood work just completed
provided the following values:
Serum Folate = 5.8 nmol/L
RBC Folate = 253 nmol/L
In addition, results indicate significant elevations in his serum methylmalonic acid concentration.
Based on the older and most recent information and data, describe and discuss his current folate and B12
status – including any/all contributing factors.
(9 points)
2013 LAB 5
Name: _______________Problem Set _______________
Lab TA: ________________________
(4) A 27 yr. old female has recently (1 week ago) returned after spending 20 months in the Swiss Alps.
She has returned early from her assignment, complaining of stress, fatigue and weight loss, and during
the last several months, had increased her smoking to about 2 packs per day (up from less than 1 pack
per day before she left). As part of her monitoring, the following data have been collected: (33 points)
HgB (g/dl)
RBC# (x 1012)
MCV [± SD] (fL)
Serum Folate (nmol/L)
24 months ago
13.6
(prior to going abroad)
16 weeks ago
12.4
8 weeks ago
10.4
4 weeks ago
8.2
4.41
91.3 [ 11.22 ]
20.4
4.07
3.72
3.44
96.8 [ 15.64 ]
100.4 [ 15.13 ]
108.7 [ 14.88 ]
7.6
6.3
5.1
Current
3.38
110.3 [ 13.52 ]
10.5
7.9

What were her Hct values at the indicated time points? (10 pts)
24 mo. ago =
16 wks ago =
8 wks. ago =
4 wks ago =
Current =
Is she currently anemic? ( Yes / No ) (1 pt)
Based on: low Hct /
low HgB / both normal (1 pt)
(1 pt) Severity =
Describe her current RBC Morphology (circle all that apply):
microcytic / normocytic / macrocytic
hypochromic / normochromic / hyperchromic
What were her RDW values at the indicated time points (10 Points)
24 mo. ago =
16 wks ago =
8 wks. ago =
4 wks ago =
Current =
Evaluate her current status taking into account all factors (8 points) (You may attach extra paper if you need
more space.)
2013 LAB 5
Name: _______________Problem Set _______________
Lab TA: ________________________
(5) You are monitoring a 53 yr. old male who began taking a drug with known Folate antagonist
(“antifolate”) properties 8 weeks ago and is now half way through his treatment regimen. (21 points)
His current lab values are:
MCV = 103.7 fL (SD = 16.3 fL)
Serum Folate = 6.5 nmol/L
RBC# = 3.94 x1012
MCH = 32.6 pg.
RBC Folate = 210.3 nmol/L
Based on these values, complete the questions below (show calulations and units)
(2 pt)What is his Hct value?
(2 pt)What is his HgB value?
(2 pt) What is his MCHC value?
(2 pt)What is his RDW value?
(2 points)Describe his current RBC Morphology (circle all that apply):
microcytic / normocytic / macrocytic
hypochromic / normochromic / hyperchromic
Is he currently anemic? ( Yes / No ) (1 pt)
Based on: low Hct /
low HgB / both normal (1 pt)
Is there evidence that he has been compliant in taking the medication with anti-folate properties? (4 pts)
Given that his treatment is scheduled to continue for 9 more weeks, should an RBC synthesis stimulating
drug (eg. Erythropoietin / ‘Procrit’) be prescribed (briefly discuss why / why not) ? (5 pts)
2013 LAB 5
Name: _______________Problem Set _______________
Lab TA: ________________________
(6) Data sets for 3 adult non-pregnant females is presented below. Match the data set to the most logical
corresponding nutritional situation as listed below.
(9 points)
Data set 1: MCV = 97.6 fL
HgB = 11.8 g/dL
RBC Folate = 256 nmol/L HoloTC = 43 pmol/L
Data set 2: MCV = 97.8 fL
MMA = 71 nmol/L
(3 pt) Situation =
ser Folate = 5.9 nmol/L  Hcyst = 17 mmol/L
HgB = 11.8 g/dL  MMA = 277 nmol/L
(3 pt) Situation =
RBC Folate = 256 nmol/L  HoloTC = 30 pmol/L ser Folate = 7.9 nmol/L Hcyst = 17 mmol/L
Data set 3: MCV = 103.4 fL  HgB = 11.8 g/dL
MMA = 71 nmol/L
RBC Folate = 378 nmol/L
ser Folate = 11.4 nmol/L Hcyst = 11 mmol/L
HoloTC = 43 pmol/L
(3 pt) Situation =
Situation A = Early B12 Deficiency
Situation B = Recovering from Folate Deficiency Anemia
Situation C = Early Folate Deficiency
Situation D = Early B12 Depletion
(7) 3 non-pregnant 35 yr.old female subjects have the following identical blood data
RBC conc. = 4.0 x 1012 /L MCV = 85 fL HgB = 13.1 g/dL Hct = 38%
(13 points)
RDW = 11.8%
Unique aspects for each subject are:
Subj. A: ser Folate = 7-8 nmol/L; RBC Folate = 340 nmol/L; (+) Antibodies against IF; smokes
Subj. B: ser Folate = 11-14 nmol/L; RBC Folate = 420 nmol/L; Kidney disease
Subj. C: ser Folate = 10-12 nmol/L; RBC Folate = 380 nmol/L; takes asthma medication
All 3 are about to start an experimental Folate deficient diet. Given the above information, answer and
briefly discuss the following questions:
Which subject would you predict to display abnormal RBC indices and/or display clinical Anemia first?
(1 pt)Subject:
(12 pts) Why?
2013 LAB 5
Name: _______________Problem Set _______________
(8) Data for 6 patients is presented below:
Patient
s Folate
(gender)
(nmol/L)
Al
(m)
Bert
(m)
Nan
(F)
Mark
(m)
Jill
(preg)
Lucy
(F)
RBC Folate
Lab TA: ________________________
(24 points)
Homocys
HgB
MCV
(nmol/L)
(mol/L)
(g/dL)
(fL)
HoloTC s B12 s MMA
(pmol/L) (pmol/L) (nmol/L)
RDW
5.7
307
14.8
14.2
93.7
43
357
104
13.4
12.5
368
18.3
8.2
108.6
29
123
284
13.4
16.8
353
8.4
9.4
103.2
44
298
86
19.2
5.8
258
17.2
13.6
96.1
44
306
93
15.8
13.2
398
7.6
8.6
84.5
42
322
124
12.1
9.1
318
12.4
13.4
91.8
32
187
237
12.2
(%)
Possible diagnosis:
Early depletion/depletion stage (Folate)
Early depletion/depletion stage (B12)
Early Deficiency stage (Folate)
Early Deficiency stage (B12)
Anemia from Folate deficiency
Anemia from B12 deficiency
Recovering from Folate Deficiency Anemia
Recovering from B12 Deficiency Anemia
Moderately Anemic (but NOT from Folate or B12 deficiency)
Based on all the above data, write the “most consistent” diagnosis (choose from diagnosis listed above)
with respect to their lab values next to the patient name. Also, indicate whether each patient should have
the following tests done. (24 Points)
Patient
Diagnosis
(from choices above)
Test for IntrinsicFactor Antibodies
(IF-Abs)
Yes or No
Test for Renal
(Kidney) Function
Yes or No
Al (m)
Bert (m)
Nan (F)
Mark (m)
Jill (Preg)
Lucy (F)
Finally, one of the 3 patients is vitamin B12 deficient. Who is it? _______________ (2 pts)
2013 LAB 5
Name: _______________Problem Set _______________
Lab TA: ________________________
(9) You are monitoring the progress of 3 patients (Bill, Greg, and Joan). Joan is suffering from a chronic
folate deficiency anemia, Bill is recovering from surgery after a traumatic motorcycle accident, while
Greg is being monitored following the accidental, prolonged ingestion (over a 5 week period) of a potent
Folate antagonist almost 2 months ago.
(29 points)
You have the data sets below. Unfortunately, the patient codes were not attached. Based on these data
sets, deduce which describes the condition and thus identify the likely patient for that data set.
Data set A:

HgB = 12.3g/dL Hct = 40.3%
(1 pt) MCV =
(3 pts) Anemia ( yes / no )
RBC# = 4.02 x1012
RDW = 19.5%
(1 pt) MCHC =
Based on: low Hct /
low HgB / both normal
RBC Morphology (circle all that apply): microcytic / normocytic / macrocytic (1 pt)
hypochromic / normochromic / hyperchromic (1 pt)
Serum Folate = 13.3 nmol/L
Serum MMA = 87 nmol/L
RBC Folate = 337 nmol/L
Serum Hcyst = 12.7 nmol/L
(2 pts) Probable patient =
------------------------------------------------------------------------------------------------------------------------------Data set B: HgB = 9.2g/dL

(1 pt) MCV =
Hct = 30.4% RBC# = 2.81 x1012
(3 pts) Anemia ( yes / no )
based on: low Hct /
RDW = 16.3%
(1 pt) MCHC =
low HgB / both normal
RBC Morphology (circle all that apply): microcytic / normocytic / macrocytic (1 pt)
hypochromic / normochromic / hyperchromic (1 pt)
Serum Folate = 4.7 nmol/L
Serum MMA = 293 nmol/L
RBC Folate = 194 nmol/L
Serum Hcyst = 19.3 nmol/L
(2 pts) Probable patient =
-------------------------------------------------------------------------------------------------------------------------------Data set C:
HgB = 10.7g/dL
(1 pt)MCV =
(3 ptS)
Anemia ( yes / no )
Hct = 31.4%
RBC# = 3.92 x1012
RDW = 12.7%
(1 pt)MCHC =
based on: low Hct /
low HgB / both normal
RBC Morphology (circle all that apply): microcytic / normocytic / macrocytic (1 pt)
hypochromic / normochromic / hyperchromic (1 pt)
2013 LAB 5
Name: _______________Problem Set _______________
Lab TA: ________________________
Ser Folate = 15.3 nmol/L
ser MMA = 71 nmol/L
RBC Folate = 457 nmol/L
ser Hcyst = 7.4 nmol/L
(2 pts) Probable patient =
(10) The 3 women below are part of a study examining a new, strict diet. At the start of the study, the
following data was collected for each woman:
(16 points)
(nmol/L)
(nmol/L)
Hct (%) HgB (g/dL) RDW (%) MCV (fL) RBC Folate Ser. Folate
(mmol/L)
Ser Homocyst.
Sally
41
13.6
12.7
92
394
15.2
11.2
Hillary
45
13.2
11.6
87
423
19.4
6.8
Louise
43
15.4
11.3
82
408
13.2
7.7
The following data was collected on Day 60 of the diet trial:
(nmol/L)
Hct (%)
(nmol/L)
HgB (g/dL) RDW (%) MCV (fL) RBC Folate Ser. Folate
(mmol/L)
Ser Homocyst.
Sally
37
12.3
16.6
99
343
12.5
17.8
Hillary
42
12.2
14.8
96
384
12.1
9.7
Louise
37
13.4
15.7
94
341
16.3
13.4
Describe any problem or risk of this new diet (4 pts)
Based on the data, which woman (if any) would you expect to become Anemic first? (1 pt)
Why? (Provide data to substantiate your answer) (3 pts)
Diet histories indicate that one of the women may have had marginal B12 status at the start of the study
and that inadequate (low) B12 intake has likely persisted through this 60 day time. Based on the data,
whichwomen is most likely to be the one with chronic, inadequate B12? (1 pt)
Why? (3 pts)
The technician doing the sample analysis informs you that one of the day 60 serum samples had evidence
of hemolysis, but is not sure which patient sample it was. Based on the above, whose day 60 sample is
the most likely to have hemolysis? (1 pt)
2013 LAB 5
Name: _______________Problem Set _______________
Lab TA: ________________________
Why? (3 pts)
(11) You are involved with a 57 yr. old female patient who presented 2 weeks ago with complaints
symptomatic for anemia. At that time, blood tests were done as indicated in the table below. Current,
follow-up blood test data is also presented. Calculate and insert the blood indice data in the table, and
answer the questions that follow. (40 points)
(12 points)
Indice
2 wks ago Current
Indice
2 wks ago
Current
RBC # (x1012/L)
3.22
3.29
MCV (fL)
HgB (g/dL)
9.2
9.0
MCH (pg)
Hct (%)
33.4
32.6
MCHC (g/dL)
RDW (%)
16.3
16.4
(3 pts) 2 wks ago: Anemia ( yes / no )
based on: low Hct /
low HgB / both normal
RBC Morphology: ( microcytic / normocytic / macrocytic ) (1 pt)
( hypochromic / normochromic / hyperchromic ) (1 pt)
(3 pts) Currently: Anemia ( yes / no )
based on: low Hct /
low HgB / both normal
RBC Morphology: ( microcytic / normocytic / macrocytic ) (1 pt)
( hypochromic / normochromic / hyperchromic ) (1 pt)
Blood tests evaluating Folate and vit.B12 were done 2 weeks ago. At that time, she was prescribed daily,
oral tablets for BOTH Folate and vit. B12. She is adamant that she has been taking the daily supplements
exactly as prescribed. Her 2 week old and current Folate and vit.B12 data are provided below. Fill-in the
evaluative data and answer the questions that follow.
(8 Points)
Current
Test
Ser. Folate (nmol/L)
2 wks ago
10.7
18.4
RBC Folate (nmol/L)
203
224
tHcy (mol/L)
17.7
16.8
Ser. B12 (pmol/L)
126
129
Status
Status
What is the evidence that she has been compliant in taking the supplements (as she claims)? (4 pts)
2013 LAB 5
Name: _______________Problem Set _______________
Lab TA: ________________________
What is your overall interpretation of her condition AND your plan for further TESTS and course of
action? (6 pts)
(12) In evaluating values and indices during the progression leading to folate deficiency anemia in an
adult male, which of the following sequences is/are correct? (2 pts)
a)
b)
c)
d)
e)
low folate intake  decrease serum folate  decreased RBC folate  an RDW = 17.7%
a prolonged RBC folate concentration of 260 nmol/L would lead to an elevated MCV
acute renal failure as well as liver disease would result in further decreases in serum Folate
all of the above
a and b only
(13) Which of the following is/are consistent with Folate deficiency anemia in an adult female? (2 pts)
a)
b)
c)
d)
e)
an MCV = 106 fL and a Hct = 30%
an MCV = 108 fL and a HgB value = 11.2 g/dL
a serum folate concentration = 5.2 nmol/L and an RBC Folate conc. = 213 nmol/L
all of the above
a and b only
(14) Concerning the inter-relationships and distinctions between folate and vitamin B12, which of the
following statements is/are true? (2 pts)
a) a lack or low amount of B12 can result in a functional folate deficiency because the folate becomes
metabolically “trapped” in an inactive form
b) the anemia blood cell indices resulting from either a folate or B12 deficiency can present identically
c) measurement of methylmalonic acid can establish that B12 is the main (primary) nutrient underlying
the deficiency if the MMA level is <100 pmol/L
d) all of the above
e) a and b only
(15) Results from a blood smear analysis state a finding of anisocytosis with hypochromacia. Based on
these observations, which of the following indice values would you expect? (2 pts)
a)
b)
c)
d)
e)
an RDW of 25.2% with an MCHC = 38 g/dL
an MCV = 78 fL and HgB = 10 g/dL
an RDW of 20.4% with an MCHC = 27 g/dl
a and b only
none of the above