HemoVoid™ Reference Applications
Human Red Blood Cells (RBC)
Bollenbach, Alexander, et al. "GC-MS and
LC-MS/MS pilot studies on the guanidine (NG)-dimethylation in native,
asymmetrically and symmetrically NG-dimethylated arginine-vasopressin peptides
and proteins in human red blood cells."
Chromatography B (2020):
Previous studies showed that human
red blood cells are rich in large (> 50 kDa) asymmetric dimethylarginine -containing
proteins of unknown identity. The study aimed to report the identity,
biological activity and concentration of NG-methylated proteins
by using GC-MS and LCMS/MS approaches. The article states “we included in our
method the use of HemoVoid™ to remove specifically most erythrocytic hemoglobin
and to improve the SDS-PAGE separation of proteins for further processing. The
HemoVoid™, … allowed removal of erythrocytic hemoglobin to a large extent from
the hemolysate. … removal of hemoglobin by this technique enabled an effective
separation by SDS-PAGE and isolation of bands, presumably by avoiding
overloading of the gels by hemoglobin.”.
Kitao, Akihito, et al. "Band 3 ectopic expression in
colorectal cancer induces an increase in erythrocyte membrane-bound IgG and may
cause immune-related anemia." International Journal of Hematology (2020):
Autoimmune hemolytic anemia (AIHA) is a rare comorbidity in
colorectal cancer (CRC) and has an unknown etiology. To better understand
cancer-related anemia, the authors’ investigated ectopic band 3 expression and
erythrocyte membrane-bound IgG in a CRC cohort. To reduce the interference from
Hemoglobin, the article states “Erythrocytes were lysed … and hemoglobin was
depleted using HemoVoid (Biotech Support Group, NJ, USA, Cat. No.
Rosin-Arbesfeld, Rina, and Ronen
SIMAN-TOV. "Article of manufacture and methods for
increasing survival of red blood cells." U.S. Patent Application
patent application describes an ex - vivo method of increasing survival of red
blood cells (RBCs). The method comprises contacting the RBCs with an activator
of the non - canonical Wnt pathway, which results in actin polymerization,
thereby increasing survival of RBCs. The invention’s description states “The
Haemolysates were enriched with over 95 % hemoglobin. For hemoglobin depletion,
the hemoglobin depletion kit of HemoVoid … was used”. Upon depletion of
hemoglobin, a reduction in cytoplasmic actin levels was observed.
Nemkov, Travis, et al. "Hypoxia modulates the purine salvage pathway and decreases red blood
cell and supernatant levels of hypoxanthine during refrigerated storage." haematologica 103.2 (2018): 361-372.
The goal of this study
was to use proteomics in part to understand hypoxanthine catabolism in vivo for stored red blood cells. It is still
unclear whether accumulation of hypoxanthine in stored red blood cell units is
clinically relevant for transfused recipients. The
article states “Leukocyte-reduced
human RBC from healthy donor volunteers were washed five times in
phosphate-buffered saline prior to lysis in distilled water with sonication.
Proteomic analyses of RBC membranes and cytosols were performed…RBC cytosolic
proteins were depleted of hemoglobin using Hemovoid™ (Biotech Support Group,
Monmouth Junction, NJ, USA), prior to high-pH reversed phase fractionation”.
Miriam M., et al. "Identification of a soluble guanylate cyclase in RBCs:
preserved activity in patients with coronary artery disease." Redox Biology (2017).
In brief, the authors aimed to investigate whether RBCs carry a functional
soluble guanylate cyclase (sGC) signalling pathway and to address whether this
pathway is compromised in coronary artery disease. The article states “Using a
commercial resin (HemoVoid™), which removes hemoglobin… and allows enrichment
of soluble cytoplasmic proteins, we established a procedure that allows fast
and reliable preparation of hemoglobin-free cell lysates from as little as 1-2
ml blood. In those samples, expression and activity of the cGMP-generating sGC,
cGMP-hydrolyzing PDE5 and cGMP-transducing PKG was assessed by enzymatic assays
and Western blot analysis”.
Amélia, et al. "Evening and morning alterations in Obstructive Sleep Apnea
red blood cell proteome." Data in
Brief (2017). http://dx.doi.org/10.1016/j.dib.2017.01.005
proteomics-based evaluation of red blood cells (RBC), the authors identified
differentially abundant proteins associated with Obstructive Sleep Apnea
Syndrome (OSA). Proteome variations between various time points were assessed.
The article states “RBC cytoplasmic fraction depleted of hemoglobin, using
HemoVoid™ system, were analyzed by two-dimensional fluorescence difference gel
electrophoresis (2D-DIGE), the 2D image software-based analyzed and relevant
differentially abundant proteins identified by mass spectrometry (MS)”.
Philipp F Lange, Pitter F Huesgen, Karen Nguyen, and Christopher M
Overall. ” Annotating N termini for the
Human Proteome Project: N termini and Nα-acetylation status differentiate
stable cleaved protein species from degradation remnants in the human
erythrocyte proteome”, J. Proteome Research., Just
Accepted Manuscript • DOI: 10.1021/pr401191w • 21 Feb 2014
article describes a goal of the Chromosome-centric Human Proteome Project to
identify all human protein species. Enucleated, erythrocytes are simple yet
proteomically challenging cells due to the high hemoglobin content (about 97%
by mass) and wide dynamic range of protein concentrations that impedes protein
identification. Using a N-terminomics procedure called TAILS, the authors
identified from the HemoVoid™ treated, soluble fraction, 778 proteins were
identified, 171 of which were not represented in either the soluble
non-depleted fraction or the membrane fraction.
HemoVoid™ On Bead Digestion Application Work On RBC by Irene Granlund, Umeå University
Katja Walpurgis, Maxie Kohler, Andreas Thomas et al. Validated hemoglobin-depletion approach for red blood
cell lysate proteome analysis by means of 2D-PAGE and Orbitrap MS.Electrophoresis.2012;
Mizukawa, B., George, A., Pushkaran,
S. et al. Cooperating G6PD mutations associated with severe
neonatal hyperbilirubinemia and cholestasis.Pediatric Blood Cancer.2011;56: 840-842.
Sudha Neelam, David G Kakhniashvili,
Stephan Wilkens et al. Functional 20S proteasomes in mature human red blood
cells Experimental Biology and
RBCs in Parkinson’s Disease
Stephan, et al. "Optimizing red blood cell protein extraction for
biomarker quantitation with mass spectrometry." Analytical and Bioanalytical Chemistry (2020): 1-14.
article describes the advantage of HemoVoid™ in detection of low abundance
proteins when comparing their amounts (in percent) between four alternative extraction
conditions, stating “…
Most peptides, following HemoVoid™ extraction, showed ion abundances ranging between
1.00E+5 and 1.00E+6 (31%). In comparison to this, fewer peptides (10–23%) were within this range following extraction with all
other protocols”. With respect to potential biomarkers for Parkinson’s
Disease, the article states “For example, PRDX6 accounts for 0.4% of the total
ion abundance after DOC (deoxycholate) extraction, whereas following HV
(HemoVoid™) extraction, this increases to 8%, a 20-fold enrichment”. The
authors conclude that the HemoVoid™ method significantly reduces the
concentration of hemoglobin, resulting in an increased signal-to noise of the
remaining red cell proteins. The article describes methods to digest the
HemoVoid™ bead-bound proteome, greatly simplifying the workflow for LC-MS/MS
Elhadi, Suaad Abd, et al. "α‐Synuclein in blood cells differentiates Parkinson’s
disease from healthy controls." Annals of Clinical and Translational Neurology.
goal of this study was to determine
whether blood cells expressing α-Synuclein can differentiate Parkinson’s
disease (PD) from healthy controls. Two proteoforms
- PSer129 a-Syn (phosphorylated
pathological form in Lewy bodies) and Oxidized a-Syn levels are observed in blood cells, but both at considerably lower
concentration than total a-Syn,
so the extremely high abundance of hemoglobin interferes with their analysis.
To compensate, the article states for PSer129 α -Syn & Oxidized α -Syn
detection by immunoassay, “followed from hemoglobin clearance with HemoVoid kit
(Biotech Support Group LLC, NJ, US)”.
Red Blood Cells,
Machado, Patrícia Isabel Pires. Pyruvate kinase and
glucose-6-phosphate dehydrogenase deficiencies and their association with
malaria–population genetics and proteomic studies. Diss. Universidade do Porto, 2013.
Walpurgis, Katja, et al. "Effects of gamma irradiation and 15 days of subsequent ex
vivo storage on the cytosolic red blood cell proteome analyzed by 2D DIGE and
Orbitrap MS." PROTEOMICS-Clinical Applications (2013).
P. Falciparum Clone 3D7
Cultured In Human Erythrocytes
Lasonder E, Green JL, Camarda G, Talabani H, Holder AA, Langsley G, Alano P. The Plasmodium falciparum schizont phospho-proteome
reveals extensive phosphatidylinositol and cAMP-Protein Kinase A signalling. J Proteome Research. 2012;
Species Agnostic – Applications
in non-human samples
Puente-Marin, Sara, et al. "In Silico Functional Networks Identified in Fish Nucleated Red Blood
Cells by Means of Transcriptomic and Proteomic Profiling." Genes 9.4 (2018): 202.
Nucleated red blood
cells (RBCs) of fish have, in the last decade, been implicated in several
immune-related functions, such as antiviral response, phagocytosis or
cytokine-mediated signaling. Label-free shotgun proteomic analyses were carried
out for in silico functional pathway profiling of rainbow trout RBCs. The
article states “The
cytosolic fraction, approximately 300 μL, was depleted of hemoglobin using
HemoVoidTM kit (Biotech Support Group, Monmouth
Junction, NJ, USA), in accordance with the manufacturer’s instructions”.
Puente-Marin S, Chico V et al. Identification of diverse defense mechanisms
in trout red blood cells in response to VHSV halted viral replication [version
1; referees: awaiting peer review].
F1000Research 2017, 6:1958 (doi: 10.12688/f1000research.12985.1)
Fish nucleated red blood cells (RBCs) generate a wide variety of immune-related
gene transcripts when viruses replicate inside them and are their main target
cell. However, the objective of this study not yet explored, was to determine
the immune response and mechanisms of fish RBCs against viruses targeting other
cells or tissues. The article states “a new proteomic analysis method was carried out that combines
fractionation into cytosolic and membrane fractions, haemoglobin removal of the
cytosolic fraction, protein digestion, pH reversed-phase peptide fractionation
and finally LC ESI-MS/MS analysis of each of the fractions… . Briefly, the
haemoglobin of the cytosolic fraction was removed using a column of HemoVoid™ kit (Biotech Support Group, Monmouth Junction, NJ), following the